Integrating computational and physiological analyses of galactomannan biosynthesis in Cyamopsis tetragonoloba

Integrating computational and physiological analyses of galactomannan biosynthesis in Cyamopsis tetragonoloba | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Integrating computational and physiological analyses of galactomannan biosynthesis in Cyamopsis tetragonoloba Ruchi Singh This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7730051/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Cyamopsis tetragonoloba (guar or cluster bean) is an important arid legume valued for its medicinal properties and industrial gum production. The principal constituent of guar gum is galactomannan, a polysaccharide composed of galactose and mannose, synthesized in the seed endosperm through the galactomannan biosynthetic pathway. Despite its economic significance, little information is available on the structural and functional aspects of enzymes involved in this pathway. In the present study, key enzymes including α-D-galactoside galactohydrolase, β-1,4-mannan synthase (MS), phosphomannose isomerase (PMI), UDP-galactose 4-epimerase (UDP-G4E), and galactomannan galactosyltransferase (GGT) were characterized using bioinformatic approaches. Secondary and three-dimensional structures were predicted along with ligand-binding and active site identification. Physicochemical properties such as isoelectric point (pI), aliphatic index (AI), grand average of hydropathicity (GRAVY), instability index, hydrophobicity, and transmembrane activity were also analyzed, providing valuable insights into enzyme stability and functionality. To complement the computational analysis, guar plants were subjected to drought stress to examine physiological and gum-related responses. Relative water content, photosynthetic rate, and transpiration rate significantly decreased under water deficit, whereas anthocyanin content increased, suggesting enhanced antioxidant defense against reactive oxygen species (ROS). Pod yield was reduced under stress; however, seed gum yield and viscosity increased. Furthermore, expression of genes involved in galactomannan biosynthesis was upregulated under drought conditions, indicating a stress-induced shift towards enhanced gum accumulation. Overall, this multidisciplinary study integrates computational enzyme characterization with physiological and biochemical analyses, providing a comprehensive understanding of galactomannan biosynthesis and drought-induced gum modulation in C. tetragonoloba. These findings may facilitate targeted crop improvement strategies and industrial applications of guar gum under climate-stress scenarios. Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Introduction Cyamopsis tetragonoloba (guar or cluster bean) is a member of the Leguminosae family and has been valued since ancient times for its medicinal properties in treating microbial infections, tumors, inflammation, night blindness, and respiratory ailments (Wang and Morris 2007 ; Kumar et al. 2024 ). In addition to its medicinal value, guar is of high economic importance due to its seed endosperm, which is a rich source of gum with wide industrial applications. Guar gum has attracted considerable research attention for its thickening, stabilizing, and gelling properties in aqueous media, making it useful in food, pharmaceutical, and industrial formulations (Mudgil et al. 2014 ). The main constituent of guar gum is galactomannan, a polysaccharide consisting of a linear (1→4)-β-linked mannose backbone with (1→6)-linked α-D-galactopyranosyl side chains (Heyne and Whistler 1948 ). The mannose-to-galactose (M:G) ratio varies among species, with guar galactomannan typically having an M:G ratio of ~ 1.6 (Edwards et al. 1992 ; Dos Santos et al. 2015 ). While pure mannan is insoluble in water, increasing galactose substitution enhances solubility by extending the polymer chain (Stephen 1983 ; Noble et al. 1986 ). Functionally, galactomannans contribute to water imbibition, drought avoidance during germination, and serve as carbohydrate reserves for seedling development (Grant Reid and Derek Bewley 1979 ). In guar, galactomannan is synthesized during seed development and stored as a mucilaginous layer in the endosperm (Kučka et al. 2022 ), accounting for 26–32% of seed dry weight (Kays et al. 2006 ). Galactomannan biosynthesis is closely linked to central carbohydrate metabolism, with sucrose acting as a key connecting metabolite (Fig. 1). In plants, sucrose is transported from source leaves to sink tissues and is cleaved either by invertase into glucose and fructose or by sucrose synthase into UDP-glucose and fructose (Lammens et al. 2009 ). Glucose and fructose are phosphorylated by hexokinase to generate glucose-6-phosphate and fructose-6-phosphate, respectively. The latter is converted into mannose-6-phosphate (Man-6-P) by phosphomannose isomerase, which also functions in the reverse direction during mannose utilization (Sharma et al. 2014 ). GDP-mannose pyrophosphorylase and UDP-galactose 4-epimerase generate the activated sugar precursors GDP-D-mannose and UDP-D-galactose, respectively. Two tightly membrane-bound glycosyltransferases then catalyze polymer formation: GDP-mannose-dependent mannosyltransferase extends the β-1,4-linked mannose backbone, while UDP-galactose-dependent galactosyltransferase adds galactose residues via α-1,6 linkages (Sharma et al. 2022 ). The activities of these enzymes increase in parallel, maintaining a constant M:G ratio in the polymer. During germination, galactomannan degradation is carried out by β-mannosidase, β-mannanase, and α-galactosidase, which hydrolyze the mannan backbone, side chains, and oligomannans, respectively (Grant Reid and Meier 1973 ). Although aspects of galactomannan yield have been studied in several species such as coffee (Pré et al. 2008 ), guar (Gresta et al. 2017 ), fenugreek (Wang et al. 2012 ), and Caesalpinia pulcherrima (Cerqueira et al. 2009 ), information on the genes and proteins of the galactomannan biosynthetic pathway remains limited. Reports are available on cDNA libraries for developing seeds of guar (Gresta et al. 2017 ) and coffee (Pré et al. 2008 ), but detailed structural and functional characterization of the pathway enzymes in guar is still scarce. Previous computational studies have successfully predicted enzyme structures and functions in secondary metabolic pathways, such as monoterpenoid biosynthesis in Mentha (Sanchita et al. 2014) and Ocimum basilicum (Yilmaz and Karik 2022 ), and triterpenoid saponin biosynthesis in Centella asiatica (Kumar et al. 2013 ). In guar, the galactomannan biosynthesis pathway, gum content, and expression of biosynthetic proteins are also influenced by environmental stresses such as drought. Drought stress reduces physiological parameters including relative water content, osmotic potential, photosynthetic and transpiration rates, and pigment concentrations, while enhancing antioxidant accumulation (Pandey et al. 2022 ). Interestingly, pod yield decreases under stress, but gum yield per seed often increases, accompanied by altered expression of biosynthetic genes. The present study therefore integrates computational and physiological approaches to analyze galactomannan metabolism in C. tetragonoloba . Specifically, we focus on structural and functional prediction of key biosynthetic enzymes, their binding site analysis, and the expression of pathway genes under drought stress. In parallel, we examine the physiological performance of guar under water deficit. Together, this information may help to unravel the regulation of galactomannan biosynthesis and provide a basis for improving gum yield and quality under stress conditions. Materials and methods Sequence retrieval Sequence retrieval The sequences of five proteins of galactomannan biosynthetic pathway of Cyamopsis tetragonoloba (Accession no. P14749, ADF47159, AAX31279, CAA32772, O65781, CAI79402) were retrieved from the protein sequence repository of NCBI database ( www.ncbi.nlm.nih.gov/ ) (Table 1 ). The protein sequences were retrieved in FASTA format and used for further analysis. Table 1 Protein sequences considered for the study S. No. Protein name Accession no. Length (AA) 1 α-D-Galactoside galactohydrolase P14749.1 411 2 β-1,4-Mannan Synthase ADF47159.1 526 3 Phosphomannose isomerase AAX31279.1 428 4 UDP-Galactose 4-epimerase O65781.1 350 5 Galactomannan Galactosyltransferase CAI79402.1 435 Functional analysis The function of a protein depends on its combination of domains (Fong and Marchler-Bauer 2008 ). For functional characterization of protein the conserved domains were searched through Conserved domain database (CDD) (Marchler-Bauer et al. 2011 ). The motifs were predicted using MEME suite ( http://meme.nbcr.net/meme/cgi-bin/meme.cgi ). The parameter of motif size was set at 6 and 50 as minimum and maximum, respectively. The gene ontology classification was also done by using Uniprot database ( http://www.uniprot.org ). Each protein was classified in three classes, biological process, molecular function and cellular component. Primary Structure Analysis Primary structure analysis or physico-chemical characterization was done by using the Expasy’s ProtParam server ( https://web.expasy.org/protparam/ ). ProtParam was used to compute the biophysical and biochemical properties like isoelectric point (pI), molecular weight, instability index (Guruprasad et al. 1990 ), aliphatic index (Ikai 1980 ) and grand average hydropathy (GRAVY) (Kyte and Doolittle 1982 ). Secondary Structure Prediction CFSSP ( https://www.biogem.org/tool/chou-fasman/ ) was employed for calculating the secondary structural features present in the protein sequences of galactomannan biosynthetic pathway considered in this study. The protein sequences were analyzed to predict the contribution of alpha helix, beta sheet and random coil structures (Chou and Fasman 1974 ) {30]. Model building and evaluation The three dimensional structure of the protein was predicted by threading alignment modelling program using I-TASSER server ( https://zhanggroup.org/I-TASSER/ ) (Yang et al. 2015 ; Zheng et al. 2021 ). The templates were selected from a link connecting to PDB database ( https://www.rcsb.org/ ) consisting of known structures. The high resolution crystal structures of template proteins were considered for threading modelling. The template with maximum identity value and minimum e-score was selected for each protein sequence to build a model. The energy minimization was performed to stabilize the modelled protein with Swiss PDB Viewer ( https://spdbv.unil.ch/ ) (Johansson et al. 2012 ). The model having higher C value (Confidence score) were chosen for further analysis. Structure visualization was performed through Pymol ( https://pymol.org/2/ ). Structure validation After the model generation, its quality assessment was done, based on both geometric and energetic aspects. The stereochemical properties of obtained protein model were checked using RAMPAGE server (Lovell et al. 2003 ). The stereochemical properties of the proteins were assessed by Ramachandran plot analysis (Ramakrishnan and Ramachandran 1965 ). The validation for modelled structures was done to determine whether the residues were falling in the most favoured region in the Ramachandran plot by using PROCHECK (Laskowski et al. 1996 ). Protein ligand binding site analysis All modelled proteins were analyzed for ligand binding site by using COACH server ( https://zhanggroup.org/COACH/ ). This server generates complementary ligand binding sites using two comparative methods, TM-SITE and S-SITE, which recognizes ligand binding templates from BioLiP protein function database by binding specific substructure and sequence profile comparisons. These predictions will be combined with results from other methods including COFACTOR ( https://zhanggroup.org/COFACTOR/ ), FINDSITE (Brylinski and Skolnick 2008 ) and ConCavity ( https://compbio.cs.princeton.edu/concavity/ ) to generate final ligand binding site predictions. Active site prediction Protein-protein superposition, brings ligand into the target active site with an orientation similar to the one found in the original ligand-receptor crystal structure (Srivastava et al. 2010 ). The ligand binding site of modelled protein was predicted by superimposition of known template proteins by using COACH server ( https://zhanggroup.org/COACH/ ). The RMSD was calculated using the superimposition between matched pairs. Ligand showing higher RMSD value and Z-value more than 1 were used for docking study. Active site residues and ligand binding site residues are shown in Table 9 . Table 9 Parameters used for the 3D structure, ligand binding site and active site prediction of the pathway protein by using I-TASSER server Template used for modeling Ident 1 Ident 2 Cov Norm Z score Cluster Size C-Score Lig Name Lig binding Residues C score EC TM score RMSDa IDENa Cov EC No Active site res PDB hit for active site 1 1uasA 0.72 0.63 0.88 2.73 77 0.66 GLA 63, 98, 99,140, 148, 175, 177, 209, 211, 228, 232, 264 0.737 0.875 0.62 0.715 3.2.1.22 177,232 1uasA 2 4hg6A 0.18 0.27 0.96 1.86 37 0.36 UDP 98,99,100,102,129,165,166,169,189,190,322 0.203 0.461 5.61 0.071 2.4.1.41 NA 2d7iA 3 1pmiA 0.39 0.42 0.96 2.84 11 0.17 KOJ 55,108,117,119,144,146,282,284 0.627 0.616 2.88 0.192 5.3.1.8 291,301 1qwrA 4 4ouaB 0.09 0.20 0.93 1.47 5 0.14 OTR 38,177,178,181,193,194,195,196,199 0.117 0.334 7.77 0.029 1.4.1.13 NA 2vdcF 5 3enkA 0.56 0.54 0.97 3.85 556 0.98 NAD 10,12,13,14,15,34,35,36,37,38,39,63,64,65,86,87,88,90,105,128,129,130,154,158,182,183,184,185 0.681 0.908 1.30 0.424 5.1.3.2 14,154,183,205,238 2c20A 1) α-D-Galactoside galactohydrolase 2) β-1,4-Mannan Synthase 3) Phosphomannose isomerise 4) Galactomannan Galactosyltransferase 5) UDP-Galactose 4-epimerase Molecular docking The docking of ligand with modelled protein was performed by Swiss Dock ( http://www.swissdock.ch/ ) at ExPASY server. In this approach, the full flexible ligand was used for docking while keeping the protein in a fixed orientation in space. The negative and lower value of binding energy as well as more numbers of cluster involved showed favoured binding between ligand and target. Plant material and drought experiment Cluster bean ( Cyamopsis tetragonoloba ) seeds were grown in pots in soil and placed in growth chamber (PGC-105 HID; Percival Scientific, Inc., 505 Research Drive, Perry, IA 5022D, USA) for the study. The temperature variation was between 25–32°C and relative humidity was in the range of 50–70%. The photosynthetic photon flux density increased gradually from 6:00 to 12:00 h which ranged from (50–1300 µmol m − 2 s − 1 ) and then decreased till 7:00 pm to maintain light and dark cycle for the plants. The two months old plants were subjected to drought by withdrawal of water. Measurement of relative water content To measure relative water content of the varieties three to five mature leaves at 0, 1st, 5th, 10th day of drought and at rewatered stage were harvested early morning to measure the fresh weight. These leaf samples were kept overnight in water to measure the turgid weight (TW) the next day after they had attain turgidity and then these were kept in oven at 70°C for 72h. The dry weight was recorded and the relative water content RWC was calculated as RWC (%) = [(FW − DW)/ (TW − DW)] ×100. Measurement of the osmotic potential Osmotic potential of plants under different days of drought were determined in the leaves using a Wescor 5520 osmometer (Wescor, Logan, Utah, USA) as described by Ball and Oosterhuis (2005). Osmotic potential measurements were taken with leaf discs weighing approx 10-15mg each. Gas exchange measurements The net photosynthesis rate ( A ), stomatal conductance ( gs ), transpiration ( E ), and WUE were measured in fully expanded leaves of guar at the regular interval with portable photosynthesis system GFS-300 (Gas exchange and fluorescence measuring system). The photosynthetic photon flux density was maintained at 800 µmol m − 2 s − 1 , CO 2 level in the leaf cuvette was 400 ppm, vapour pressure deficit (VPD) level was less than 2KPa and temperature of the leaf was at 30°C. Chlorophyll estimation The photosynthetic pigments such as chlorophyll and carotenoid were measured using method of Wellburn(Wellburn 1994 ). Leaf tissue of 20–25 mg was taken each from control and drought treated plants. These were kept in 1 ml of 80% acetone in dark for pigment extraction, then OD were recorded at 470, 647 and 663 nm for chl a, chl b and carotenoids determination. Physicochemical characterization of gum A Physiochemical property of the seed was measured as follows: Sample collection Seeds were collected from pods of guar plants of both watered and drought treated conditions. Seeds were soaked in water to remove endosperm and were kept in a refrigerator at 2–4° C for at most 24 h prior to isolation and purification. Extraction and Isolation of the seed gum Extraction and isolation of gum from seeds of guar were done using the procedure of Liyanage et al 2015 (Liyanage et al. 2015 ). The seeds were soaked in water to remove the endosperm for gum extraction. These were then dried in oven to remove moisture .Approximately (10–12 g) of the endosperm was grinded to a powder and left overnight in 50ml hexane. The powder was properly washed the next day in hexane and was filtered. The filtrate obtained was dried to remove excess hexane. 100ml water was then added to this and kept overnight. The next day it was filtered and washed in water for 2–3 times. The filtrate was centrifuged at 12000rpm for 10 mins and supernatant was taken. To this supernatant add ethanol (1.5 times the supernatant). A thick substance precipitates out as gum. This precipitate was oven dried at 50 ⁰ C till it was powder. To purify the gum, the dried gum was dissolved in isopropanol (same volume as that of gum) with continuous stirring until completely dissolved and then finally dried. Determination of Specific Rotation of Gums Each gum sample (.01%) were filtered and equilibrated at 27° C of temperature. Optical rotations were determined for the three samples using polarimeter according to Dawber et al. (Dawber et al. 1988 ). The specific rotation of gum samples were measured in degrees. Viscosity Gum solutions of 0.01% were prepared by continuous stirring using a magnetic stirrer until the solution gelatinized. The actual viscosity of the gum was measured at 30° C at 6 rpm by a Well Brookfield Cone/Plate Microviscometer model R VT according to Stoloff et al (Stoloff 1958 ). The viscometer reading in centipoise (cP) was multiplied by a factor equivalent to the spindle number that has been used. Ash and carbon content Percentage ash and carbon content was determined using Analyser Multi N/C (Analytic Jena, Germany) 2100S. 40–50 mg of gum sample was used for estimation of carbon and ash content. Results and discussion Functional analysis of pathway proteins Domains are evolutionarily conserved units in proteins and frequently correspond to recurrent structural and functional units (Basu et al. 2008 ). The particular function of a protein depends on its combination of domains; 80% of eukaryotic proteins have more than one domain. The galactomannan biosynthesis pathway proteins were analyzed for the availability of conserved domains. The domain having least e-value was selected corresponding to each protein. The retrieved conserved domains and their position in proteins were analyzed. Galactomannan Galactosyltransferase has single domains indicating that they did not participate in the formation of multi-domain structure. However α-D-Galactoside galactohydrolase, β-1,4-Mannan Synthase, Phosphomannose isomerise, UDP-Galactose 4-epimerase has two or three domains and participated in multiple domain structure (Table 2 ).Three motifs for each conserved domains were predicted. The motifs might be involved in binding site formation (Sanchita et al. 2013). The substrates of these proteins bind at these motifs of proteins (e.g enzymes). The motifs are represented in form of regular expression and web logo that explains the frequency of occurrence of residues of any motif at minimum 2 sites (Table 3 ). Table 2 Conserved Domain found in pathway proteins Protein Conserved Domain Position in the protein E-value α-D-Galactoside galactohydrolase GH27 56–319 2.67e − 159 PLN02692 1-411 0e + 00 β-1,4-Mannan Synthase CESA_CaSu_A2 91–327 1.4e − 137 Glyco_tranf_2_3 91–327 1.51e − 25 Phosphomannose isomerase PLN02288 14–407 0e + 00 Man A 12–404 5.55e − 55 UDP-Galactose 4-epimerase UDP_G4E_1_SDR_e 5-334 0e + 00 PLN02240 1-349 0e + 00 Galactomannan Galactosyltransferase PLN03181 1-435 0e + 00 Table 3 Domains with corresponding motifs and their regular expression Protein No. of Motifs Regular expression Logo E-value Width Site count R e entropy α-D-Galactoside galactohydrolase 3 [RQ][YT][PP][MG] 3.3e + 000 6 2 25.6 [WR][SN][HF][SG][IC] 9.8e + 000 6 2 24.2 [KF][YF][SD][NM][CE] 1.2e + 001 6 2 24.6 β-1,4-Mannan Synthase 3 A[IT][YKF]D[AL][LGD][FG][WQG][PKH]D[RHA][TFD][FT] 4.1e + 000 3 13 44.1 Q[HA]RW[SK][FC] 1.3e + 001 2 6 22.9 [SH][LS[TRID]Y[ASH][FW] 5.3e + 001 4 6 20.4 Phosphomannose isomerase 3 CS[VM][LK][TN]Y[KD][WQ]G 1.5e + 000 2 9 36.3 E[IC][ID][RE]C[MI] 3.8e + 000 2 6 25.6 [SI]Q[FA][HPD] 3.7e + 001 2 6 23.9 UDP-Galactose 4-epimerase 3 I[GED]E[MID][CP]R[DG][IQ][YWP][NR][WNA] 8.4e − 002 3 11 39.6 E[QL][NE][WK][IA] 6.7e + 001 2 6 23.8 [QP][QL]V[MA][VA][GRRP] 8.1e + 001 2 9 34.2 Galactomannan Galactosyltransferase 3 I[TL][LH][VF]TG[SC][QP][KC][RN][GC]H 7.2e − 002 2 13 51.9 D[VC]W[VN][SG]M 1.6e + 001 2 6 24.6 H[SG]W[TE][GL] 6.0e + 001 2 6 23.8 Gene ontology classification of pathway proteins The gene ontological classification of the proteins was analyzed in terms of biological process, molecular function and cellular component (Table 4 ). α-D-Galactoside galactohydrolase, Phosphomannose isomerise, UDP-Galactose 4-epimerase, enzymes involved in the carbohydrate metabolism while β-1,4-Mannan Synthase, Galactomannan Galactosyltransferase are the membrane bound enzymes and participated in cell wall organisation, biogenesis and degradation process. Phosphomannose isomerise, mannan synthase participated in mannan biosynthesis but PMI also involved in Zn ion binding. UDP-Galactose 4-epimerase catalyzes the formation of UDP galactose from UDP-glucose or fructose. Table 4 Gene ontology classification of enzymes is dividing into three group biological process, molecular function and cellular component . Protein Biological Process Molecular Function Cellular Component 1 α-D-Galactoside galactohydrolase Carbohydrate metabolic process, catalytic and enzymatic activity, α galactosidase activity and raffinose α galactosidase activity Glycosidase, Hydrolase 2 β-1,4-Mannan Synthase Cell wall biogenesis and degradation, metabolic process and cell wall organization, GDP-mannose + (mannan)(n) = GDP + (mannan)(n + 1). Transferase activity, transferring glycosyl groups Golgi apparatus, and integral component of membrane 3 Phosphomannose isomerase Carbohydrate metabolic process, GDP mannose biosynthetic process Mannose-6-phosphate isomerase activity, zinc ion binding 5 UDP-Galactose 4-epimerase Carbohydrate metabolic process, galactose metabolic process, Ligand NAD Coenzyme binding, UDP-glucose 4-epimerase activity, Isomerase activity 6 Galactomannan Galactosyltransferase Metabolic process, cell wall organisation Transferase activity, transferring hexosyl groups Golgi apparatus, and integral component of membrane Physicochemical properties of proteins Parameters computed using Expasy’s ProtParam tool given in Table 5 . The calculated isoelectric point (pI) is useful because at pI, solubility is least and mobility in an electro focusing system is zero. Isoelectric point (pI) is the pH at which the surface of protein is covered with charge but net charge of protein is zero. At pI proteins are stable and compact. The computed pI value of all protein except α-D-Galactoside galactohydrolase is more than 7 indicating the net electrical charge of proteins is zero at basic pH value. However, α-D-Galactoside galactohydrolase showed less than 7 (pI < 7) reveals that net electrical charge is zero at acidic pH value. The computed isoelectric point (pI) will also be useful for developing buffer system for purification by isoelectric focusing method. The instability index provides an estimate of the stability of protein in a test tube. This method assigns a weight value of instability. Using these weight values, it is possible to compute an instability index. A protein whose instability index is smaller than 40 is predicted as stable, a value above 40 predicts that the protein may be unstable (Guruprasad et al. 1990 ). The instability index value for these proteins was found to be ranging from 28.57 to 40.36. β-1, 4-Mannan Synthase and Phosphomannose isomerise showed comparatively high instability index 40. The very high aliphatic index of protein sequences indicates that β-1, 4-Mannan Synthase and Phosphomannose isomerise proteins may be stable for a wide temperature range. The lower thermal stability of α-D-Galactoside galactohydrolase, UDP-Galactose 4-epimerase and Galactomannan Galactosyltransferase was indicative of a more flexible structure when compared to other protein. The Grand Average Hydropathy (GRAVY) value for a protein is calculated as the sum of hydropathy values of all amino acids, divided by the number of residues in the sequence. GRAVY indices were found in the range from − 0.592 to 0.106. This low range of value indicates the possibility of better interaction with water. Negative value of hydropathy showed hydrophilic proteins while positive showed hydrophobic nature of the protein (Kyte and Doolittle 1982 ). Except β-1, 4-Mannan synthase all studied proteins showed negative value of hydropathy and these are hydrophilic nature towards water. Table 5 Physicochemical parameter of pathway proteins is computed by using Expasy’s ProtParam tool. Protein AA Mol wt pI Instability Index Aliphatic Index GRAVY α-D-Galactoside galactohydrolase 411 45135.5 4.97 32.11 76.69 -0.359 β-1,4-Mannan Synthase 526 60677.4 9.28 40.35 100.78 0.106 Phosphomannose isomerase 428 47773.4 6.60 40.36 89.72 -0.223 UDP-Galactose 4-epimerase 350 38373.7 6.72 28.57 87.74 -0.198 Galactomannan Galactosyltransferase 435 50956.0 7.98 35.57 72.44 -0.592 Structural analysis of modelled pathway proteins The secondary structure of proteins was analyzed for considering relative availability of alpha helix, extended strand and random coils. The percentage distribution of predicted secondary structure features are represented in Table 6 . The results revealed that the predicted alpha helix dominated among secondary structure elements followed by extended strand, random coils for all protein sequences. Present study findings were similar to the earlier results reported in Withania somnifera (Sanchita et al. 2014). Table 6 Secondary structure elements identified in proteins. Protein Alpha Helix (%) Extended strand (%) Random Coil (%) α-D-Galactoside galactohydrolase 62.5 59.6 14.6 β-1,4-Mannan Synthase 85.7 53.8 9.5 Phosphomannose isomerase 82.2 67.3 13.1 UDP-Galactose 4-epimerase 68.0 38.6 12.9 Galactomannan Galactosyltransferase 62.5 61.1 13.1 Three-dimensional structures were predicted for proteins where such data is unavailable. There is lack of experimental structures for the proteins considered. The modelling of the three dimensional structure of these proteins were performed by threading alignment web server, I-TASSER. The template having maximum identity (maximum alignment coverage) and highest c value was selected and models were created using I-TASSER server. The model having lowest energy was considered for further study (Fig. 2). The stereochemical quality and accuracy of the predicted models were evaluated using Ramachandran map calculations computed with the PROCHECK program. The result revealed that the modelled structure for all protein sequences have 76.5%, 72.7%, 74.3%, 76.5%, 80.9% and 56.2% residues for α-D-Galactoside galactohydrolase, β-1,4-Mannan Synthase, Phosphomannose isomerise, UDP-Galactose 4-epimerase and Galactomannan Galactosyltransferase respectively in most favoured region (Table 7 ). The identified Ramachandran plots represent a good quality of predicted models. The structures were naturally stable with low energy value. The RMSD values identified through superimposition between selected template structure and modelled structure. Table 7 Percentage distribution of amino acid residues in ramachandran plot. Protein Residues in most favored region (%) Residues in additionally allowed region (%) Residues in generously allowed region Residues in disallowed region α-D-Galactoside galactohydrolase 76.5 19.0 3.1 1.4 β-1,4-Mannan Synthase 72.7 19.5 4.6 3.1 Phosphomannose isomerase 74.3 21.7 2.1 1.9 UDP-Galactose 4-epimerase 80.9 15.7 3.0 0.3 Galactomannan Galactosyltransferase 56.2 32.8 7.3 3.6 Ligand binding site prediction for modelled proteins Modelled proteins of galactomannan biosynthetic pathway were analyzed for ligand binding site prediction. PMI showed minimum number of ligand binding residues while UDP-G4E has maximum. α-D-Galactoside, MS, PMI, GT, UDP-G4E have shown GLA, UDP, Zn, OTR and NAD as probable ligands, respectively (Fig. 3). All modelled proteins showed more than 88% sequence coverage of templates. GT have minimum clusters size while UDP-G4E showed maximum templates in a cluster (Table 8 ). Table 8 Trans-membrane activity of pathway proteins No of transmembrane helices Start from End to Helices length Score Orientation α-D-Galactoside galactohydrolase 1 124 143 20 2255 o-i β-1,4-Mannan Synthase 4 78 100 23 1901 o-i 409 428 20 25554 i-o 438 467 30 771 o-i 548 571 20 2160 i-o Phosphomannose isomerase 2 303 322 20 832 o-i 413 432 20 630 i-o UDP-Galactose 4-epimerase 2 50 69 20 516 i-o 112 133 22 752 o-i Galactomannan Galactosyltransferase 2 23 43 21 527 i-o 77 98 22 2531 i-o Active site prediction in the modelled proteins All modelled proteins were also analyzed for active sites (Fig. 4). MS, PMI and GT did not show any active site residue while α-D-Galactoside and UDP-G4E have 2 and 5 active site residues, respectively (Table 8 ). MS, PMI and GT also showed less c value for the active site prediction. These are membrane proteins and for membrane proteins c value is always less. C Score EC is the confidence score for the EC number prediction. C Score EC values range between 0–1, where a higher score indicates a more reliable EC number prediction. TM score is a measure of global structure similarity between query and template protein. RMSD a is the RMSD between residues that are structurally aligned by TM align. IDEN a is the percentage sequence identity in the structurally aligned region. Docking study of pathway protein with their ligands Molecular docking can fit molecules together with a favorable configuration to form a complex system. The structural information from the theoretical modeled complex can help us to understand the catalytic mechanism of an enzyme. The modelled pathway proteins were studied for docking with their probable ligand respectively. UDP-G4E showed minimum enthalpy energy and fitness energy thus it forms more stable UDP galactose which further converted in to galactomannans (Table 9 ). However, PMI showed comparatively higher enthalpy change and catalyze reversible reaction and forms less stable compound. Ligand binding at appropriate configuration (with lowest enthalpy change) is given in Fig. 5. Physiological performance of the guar plants under drought stress Milan variety of guar was imposed to drought for 10 days. Relative water content of the leaf dropped up to 50% at the 10th day of drought (DD) (Fig. 6A). Osmotic potent also decreased under the drought and plants were not fully recovered after the one week of recovery (Fig. 7B). The photosynthesis rate ( A ) was decreased by 66.2% in 10DD, however stomatal conductance ( g s ) and transpiration rate ( E ) did not change significantly under drought stress (Fig. 7A, B, C). Water use efficiency ( WUE ) decreased by 61% under drought stress. However, fully recovered in one of recovery (Fig. 7D). The chlorophyll content decreased up to 60% in comparison to watered plants. Chlorophyll a/b ratio did not changed significantly under drought stress (Fig. 8A, B). carotenoid content increased dramatically at the 5th DD and did not recovered after the recovery period (Fig. 8C). Anthocyanin content increased upto 80 µmolper gram of fresh weight. Further it decreased after the rewatering of one week (Fig. 8D). Drought causes decrease in water content which is accompanied by loss of turgor and wilting, closure of stomata, reduction in photosynthesis, and interference with many other basic metabolic processes (Singh et al. 2014 ). Drought stress primarily decreases the photosynthesis rate due to decrease in internal CO 2 concentration resulting from stomatal closure (Chaves et al. 2009 ; Vishnyakova et al. 2023 ). Chlorophyll and carotenoid are light harvesting photosynthetic pigments which showed changes to avoid the inhibition in photosynthesis reaction centers under drought stress. These pigments also degrade which leads to deterioration of thyllakoid membranes (Munné-Bosch and Alegre 2000 ). Carotenoids act as accessory light harvesting pigments thus transferring light energy to chlorophylls. Anthocyanins are another class of pigments which also have photoprotective effect. Under stress conditions to avoid photooxidative damage to plant tissues anthocyanins functions as antioxidants by scavenging reactive oxygen radicals (Ranjan et al. 2014 ). These act as light attenuators by not transferring the absorbed light energy to the photosynthetic machinery of the plants thus protecting underlying cells from high irradiance (Hogewoning et al. 2012 ; Zhu et al. 2016 ). Our study revealed that the anthocyanin content is found to increase under drought which is a protective mechanism of plant against stress. Gene expression of the proteins involved in the galactomannan biosynthesis Fru-6-P converts into Man-6-P by phosphomannose isomerise (PMI) which also functions in the reverse direction for the utilization of mannose. Expression of PMI is decreased by 15.8 fold in comparison to watered plants (Fig. 9). UDP-G4E is the enzyme which forms the direct precursor for galactomannan biosynthesis, UDP-D-galactose. In this study expression value was upregulated by 5.8 fold in comparison to control plants (Fig. 9). UDP-galactose-dependent GT transfers a galactose residue through a α-linkage to a mannose near the nonreducing end of the growing mannan chain. Expression of GT is upregulated by 20 fold while MS increased by 60.5 fold in comparison to watered plans (Fig. 9). Galactosidase is a enzyme which hydrolyse the galactose residue of galactomannan. Expression of galactosidase is down regulated by -30.6 fold in comparison to control plants (Fig. 9). Overall enzymes involved in the galactomannan biosynthesis are upregulated. However, enzymes associated with hydrolysis are downregulated. Formation of seed and fruit influenced by various metabolic processes occurred in leaves and translocated in to seeds. Drought stress affect the physiological and biochemical process which involved in the grain filling and grain quality (Sehgal et al. 2018 ; Boote et al. 2023 ). Although seed yield were decreased under water stress but seed gum yield increased under drought stress. Effect of drought on gum yield and its physiochemical properties The physiochemical properties of seed gum were studied. The yield of gum was increased with 1.9 fold under drought stress (Table 11 ). The specific rotation of .01% sample was decreased by 1.1 fold under drought stress. Drought stress increased relative conductivity of gum samples. Percentage of ash in gum is believed to be the measure of mineral quantities in the guar seed (Dakia et al. 2008 ; Zammel et al. 2021 ). The percentage of ash showed significant decrease by 2.2 fold under water stress. Measurements of viscosity showed increased by 1.2 fold (Table 11 ). The specific rotation of organic molecules such as carbohydrates is related to their structure and is a characteristic property of the substance (Sathyanarayana and Stevens 1987 ). The specific rotation of gum is thus considered as the most important criterion of purity and qualitative analysis. The percentage ash content indicates the presence of inorganic elements and mineral quantities in gum. The viscosity of any liquid represents its resistance to shearing and stirring. It is an important analytical parameter which depends on shape and size of the macromolecule (Anderson 1978 ). Viscosity is the most characteristic feature of guar gum it is due to its property to hydrate in aqueous solutions. Viscosity of the gum depends on several factors such as temperature, time, pH, concentration and ionic strength and type of agitation (Mudgil et al. 2014 ). It was observed that drought increased viscosity of the gum solutions. This property of viscosity is due to galactomannan which may increase during drought stress (Hoppe and Goswami 1999 ). Table 11 Gum yield and rheological properties of seed gum Parameter Control Drought Gum Yield (%) 8.67 ± 1.53 15.87 ± 1.87 Optical Rotation (µS cm − 1 ) 60.51 ± 3.89 54.78 ± 4.23 Viscosity (cP) 25.89 ± 3.78 32.45 ± 2.67 Carbon content (%) 34.75 ± 2.35 39.56 ± 33.12 Conclusion Cyamopsis tetragonoloba is an important legume crop used for medicine and seed gum. Seed gum is made up of galactomannan which is a polysaccharide. It is synthesized by a divert pathway from the central carbohydrate metabolism. Drought stress affects physiological and biochemical metabolism. It diverted carbon metabolism towards sink tissue. Relative water content, photosynthesis rate, transpiration rate, chlorophyll content decreased under water stress. However carotenoid and anthocyanin content increased during drought. These act as light attenuators by not transferring the absorbed light energy to the photosynthetic machinery of the plants thus protecting underlying cells from high irradiance. α-D-Galactoside galactohydrolase, β-1,4-Mannan Synthase (MS), Phosphomannose isomerise (PMI), UDP-Galactose 4-epimerase (UDP-G4E) and Galactomannan Galactosyltransferase (GGT) are the proteins involved in galactomannan biosynthesis. Therefore these proteins are studied in silico and gene expression analysis. Three-dimensional structures, ligand binding site, active sites were also predicted for these pathway enzymes. The physico-chemical properties such as pI, AI, GRAVY, instability index, hydrophobicity and transmembrane activity were also studied. The current information will provide a platform to know the structural and functional attributes of these proteins for further studies. Table 10 Docking parameters for pathway proteins Name of protein Cluster Element Full fitness (Kcal/mol) Estimated ∆G (Kcal/mol) α-D-Galactoside galactohydrolase 37 0 -1231.78 -6.06 β-1,4-Mannan Synthase 32 0 -936.42 -10.25 Phosphomannose isomerase 38 0 -1689.90 -5.23 UDP-Galactose 4-epimerase 30 4 -1929.91 -11.17 Galactomannan Galactosyltransferase 8 2 -1212.81 -6.19 Declarations Conflict of interest The author declares that no conflicting interests exist. Funding: Research in CSIR-CIMAP laboratory was supported by HRDG CSIR New Delhi, India (Grant No. Nehru PDF/LS/ EMR I/04/2017). RS is grateful to CSIR, New Delhi, India for Nehru Postdoctoral Fellowship. Author would like to thanks the Director CSIR-NBRI for allowing taking all the physiological measurements in Plant physiology lab of CSIR-NBRI. 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Tree Physiol 36:1092–1104. https://doi.org/10.1093/treephys/tpw047 Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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20:07:36","extension":"xml","order_by":36,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":181035,"visible":true,"origin":"","legend":"","description":"","filename":"35023a224208425ba8c949d50ca8bd511structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/2b7327a5198e09d5082b1bf9.xml"},{"id":93717384,"identity":"82734977-350f-411c-9a8e-730f3d260a52","added_by":"auto","created_at":"2025-10-16 20:15:36","extension":"html","order_by":37,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":187714,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/5344eaf89348b5411eb73fc8.html"},{"id":93717372,"identity":"d95cf95c-4e79-4ebd-84f1-f3abdc810573","added_by":"auto","created_at":"2025-10-16 20:15:35","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":134469,"visible":true,"origin":"","legend":"\u003cp\u003eSchematic representation of galactomannan biosynthesis.\u003c/p\u003e","description":"","filename":"Slide1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/68613b278ea712308d3cd1a0.jpg"},{"id":93717373,"identity":"a49ca4bd-caba-4893-92ef-f8cedf946370","added_by":"auto","created_at":"2025-10-16 20:15:35","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":163963,"visible":true,"origin":"","legend":"\u003cp\u003eThree-dimensional structure of the modelled proteins of galactomannan biosynthesis\u003c/p\u003e","description":"","filename":"Slide2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/91a0b111cde0ac7dc9b47350.jpg"},{"id":93716933,"identity":"be059838-6ac3-4d56-81b1-b413272f5b9a","added_by":"auto","created_at":"2025-10-16 20:07:35","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":186267,"visible":true,"origin":"","legend":"\u003cp\u003ePredicted ligand binding sites in the modelled proteins of galactomannan biosynthesis\u003c/p\u003e","description":"","filename":"Slide3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/0b7ad5d406b89e1bc0e5c61e.jpg"},{"id":93717527,"identity":"e95f0f5c-5f98-4be0-aa93-908eddfcb814","added_by":"auto","created_at":"2025-10-16 20:23:35","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":149597,"visible":true,"origin":"","legend":"\u003cp\u003ePredicted active sites in the modelled proteins of galactomannan biosynthesis\u003c/p\u003e","description":"","filename":"Slide4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/32fec39aed3b8a9311fd10da.jpg"},{"id":93717374,"identity":"92e2f04f-99ec-41a9-a2cd-8f50496404f4","added_by":"auto","created_at":"2025-10-16 20:15:35","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":188096,"visible":true,"origin":"","legend":"\u003cp\u003eDocking of proteins of galactomannan biosynthesis with corresponding ligands\u003c/p\u003e","description":"","filename":"Slide5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/af16028df90a2aa191676473.jpg"},{"id":93716944,"identity":"da0d100e-c149-4af4-bf40-9f418f58ec1f","added_by":"auto","created_at":"2025-10-16 20:07:35","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":104061,"visible":true,"origin":"","legend":"\u003cp\u003eRelative water content, Day RWC (%) (A). Predawn leaf water potential (B) measured in the leaves of \u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e under irrigated and water-deficit conditions on the 1st, 5th, and 10th day of drought stress (B). Each bar represents ±SD of five measurements\u003c/p\u003e","description":"","filename":"Slide6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/286af058a5106eaf349a3ee2.jpg"},{"id":93716938,"identity":"35b78a22-588e-45af-95f8-d6017aef769a","added_by":"auto","created_at":"2025-10-16 20:07:35","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":175033,"visible":true,"origin":"","legend":"\u003cp\u003eEffect of drought stress on net photosynthesis (A), stomatal conductance (B), transpiration rate (C), and water use efficiency (WUE) (D) in \u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e on the 1\u003csup\u003est\u003c/sup\u003e, 5th and 10\u003csup\u003eth\u003c/sup\u003e day of drought stress. Data represent the means±SD of five separate measurements.\u003c/p\u003e","description":"","filename":"Slide7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/93a04138eb0355900337c5dc.jpg"},{"id":93717528,"identity":"111c49fa-e8c3-48bc-9217-66b0a6339330","added_by":"auto","created_at":"2025-10-16 20:23:35","extension":"jpg","order_by":8,"title":"Figure 8","display":"","copyAsset":false,"role":"figure","size":209536,"visible":true,"origin":"","legend":"\u003cp\u003eChlorophyll a (A) chlorophyll a/b, carotenoid (C) and anthocyanin (D) content of \u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e on the 1\u003csup\u003est\u003c/sup\u003e, 5th and 10\u003csup\u003eth\u003c/sup\u003e day of drought stress. Data represent the means±SD of five separate measurements.\u003c/p\u003e","description":"","filename":"Slide8.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/460b0662b05b6af2beb54c66.jpg"},{"id":93716941,"identity":"60a9143a-aa13-4661-8828-d96cf785c8fc","added_by":"auto","created_at":"2025-10-16 20:07:35","extension":"jpg","order_by":9,"title":"Figure 9","display":"","copyAsset":false,"role":"figure","size":34703,"visible":true,"origin":"","legend":"\u003cp\u003eResponse of gene expression under drought stress in α-D-Galactoside galactohydrolase, β-1,4-Mannan Synthase (MS), Phosphomannose isomerise (PMI), UDP-Galactose 4-epimerase (UDP-G4E) and Galactomannan Galactosyltransferase (GGT) under control and the 10\u003csup\u003eth\u003c/sup\u003e day of drought condition in \u003cem\u003eCympopsis tetragonoloba\u003c/em\u003e. Results are presented as the ratio of expression of each gene (relative to that of ubiquitin) in drought-treated plants compared to watered controls, transformed in log10. Error bars indicate the SD of the mean (n=3)\u003c/p\u003e","description":"","filename":"Slide9.jpg","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/1ac39459d673a23a6fb16383.jpg"},{"id":105034960,"identity":"b500d7ee-cdbd-4383-84f7-c166347f087f","added_by":"auto","created_at":"2026-03-20 07:25:00","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":3045624,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7730051/v1/6432b99f-4151-45b7-bb62-6032ca9f55e5.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Integrating computational and physiological analyses of galactomannan biosynthesis in Cyamopsis tetragonoloba","fulltext":[{"header":"Introduction","content":"\u003cp\u003e\u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e (guar or cluster bean) is a member of the Leguminosae family and has been valued since ancient times for its medicinal properties in treating microbial infections, tumors, inflammation, night blindness, and respiratory ailments (Wang and Morris \u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e2007\u003c/span\u003e; Kumar et al. \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e2024\u003c/span\u003e). In addition to its medicinal value, guar is of high economic importance due to its seed endosperm, which is a rich source of gum with wide industrial applications. Guar gum has attracted considerable research attention for its thickening, stabilizing, and gelling properties in aqueous media, making it useful in food, pharmaceutical, and industrial formulations (Mudgil et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). The main constituent of guar gum is galactomannan, a polysaccharide consisting of a linear (1\u0026rarr;4)-β-linked mannose backbone with (1\u0026rarr;6)-linked α-D-galactopyranosyl side chains (Heyne and Whistler \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e1948\u003c/span\u003e). The mannose-to-galactose (M:G) ratio varies among species, with guar galactomannan typically having an M:G ratio of ~\u0026thinsp;1.6 (Edwards et al. \u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e1992\u003c/span\u003e; Dos Santos et al. \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). While pure mannan is insoluble in water, increasing galactose substitution enhances solubility by extending the polymer chain (Stephen \u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e1983\u003c/span\u003e; Noble et al. \u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e1986\u003c/span\u003e). Functionally, galactomannans contribute to water imbibition, drought avoidance during germination, and serve as carbohydrate reserves for seedling development (Grant Reid and Derek Bewley \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e1979\u003c/span\u003e). In guar, galactomannan is synthesized during seed development and stored as a mucilaginous layer in the endosperm (Kučka et al. \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), accounting for 26\u0026ndash;32% of seed dry weight (Kays et al. \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e2006\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eGalactomannan biosynthesis is closely linked to central carbohydrate metabolism, with sucrose acting as a key connecting metabolite (Fig.\u0026nbsp;1). In plants, sucrose is transported from source leaves to sink tissues and is cleaved either by invertase into glucose and fructose or by sucrose synthase into UDP-glucose and fructose (Lammens et al. \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e2009\u003c/span\u003e). Glucose and fructose are phosphorylated by hexokinase to generate glucose-6-phosphate and fructose-6-phosphate, respectively. The latter is converted into mannose-6-phosphate (Man-6-P) by phosphomannose isomerase, which also functions in the reverse direction during mannose utilization (Sharma et al. \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). GDP-mannose pyrophosphorylase and UDP-galactose 4-epimerase generate the activated sugar precursors GDP-D-mannose and UDP-D-galactose, respectively. Two tightly membrane-bound glycosyltransferases then catalyze polymer formation: GDP-mannose-dependent mannosyltransferase extends the β-1,4-linked mannose backbone, while UDP-galactose-dependent galactosyltransferase adds galactose residues via α-1,6 linkages (Sharma et al. \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). The activities of these enzymes increase in parallel, maintaining a constant M:G ratio in the polymer. During germination, galactomannan degradation is carried out by β-mannosidase, β-mannanase, and α-galactosidase, which hydrolyze the mannan backbone, side chains, and oligomannans, respectively (Grant Reid and Meier \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e1973\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eAlthough aspects of galactomannan yield have been studied in several species such as coffee (Pr\u0026eacute; et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2008\u003c/span\u003e), guar (Gresta et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2017\u003c/span\u003e), fenugreek (Wang et al. \u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e2012\u003c/span\u003e), and \u003cem\u003eCaesalpinia pulcherrima\u003c/em\u003e (Cerqueira et al. \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e2009\u003c/span\u003e), information on the genes and proteins of the galactomannan biosynthetic pathway remains limited. Reports are available on cDNA libraries for developing seeds of guar (Gresta et al. \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e2017\u003c/span\u003e) and coffee (Pr\u0026eacute; et al. \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e2008\u003c/span\u003e), but detailed structural and functional characterization of the pathway enzymes in guar is still scarce. Previous computational studies have successfully predicted enzyme structures and functions in secondary metabolic pathways, such as monoterpenoid biosynthesis in Mentha (Sanchita et al. 2014) and \u003cem\u003eOcimum basilicum\u003c/em\u003e (Yilmaz and Karik \u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e2022\u003c/span\u003e), and triterpenoid saponin biosynthesis in \u003cem\u003eCentella asiatica\u003c/em\u003e (Kumar et al. \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e2013\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn guar, the galactomannan biosynthesis pathway, gum content, and expression of biosynthetic proteins are also influenced by environmental stresses such as drought. Drought stress reduces physiological parameters including relative water content, osmotic potential, photosynthetic and transpiration rates, and pigment concentrations, while enhancing antioxidant accumulation (Pandey et al. \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e2022\u003c/span\u003e). Interestingly, pod yield decreases under stress, but gum yield per seed often increases, accompanied by altered expression of biosynthetic genes.\u003c/p\u003e\u003cp\u003eThe present study therefore integrates computational and physiological approaches to analyze galactomannan metabolism in \u003cem\u003eC. tetragonoloba\u003c/em\u003e. Specifically, we focus on structural and functional prediction of key biosynthetic enzymes, their binding site analysis, and the expression of pathway genes under drought stress. In parallel, we examine the physiological performance of guar under water deficit. Together, this information may help to unravel the regulation of galactomannan biosynthesis and provide a basis for improving gum yield and quality under stress conditions.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eSequence retrieval\u003c/h2\u003e\u003cdiv id=\"Sec4\" class=\"Section3\"\u003e\u003ch2\u003eSequence retrieval\u003c/h2\u003e\u003cp\u003eThe sequences of five proteins of galactomannan biosynthetic pathway of \u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e (Accession no. P14749, ADF47159, AAX31279, CAA32772, O65781, CAI79402) were retrieved from the protein sequence repository of NCBI database (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e\u003ca href=\"http://www.ncbi.nlm.nih.gov/\" target=\"_blank\"\u003ewww.ncbi.nlm.nih.gov/\u003c/a\u003e\u003c/span\u003e\u003cspan address=\"http://www.ncbi.nlm.nih.gov/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The protein sequences were retrieved in FASTA format and used for further analysis.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eProtein sequences considered for the study\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eS. No.\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eProtein name\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eAccession no.\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eLength (AA)\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eP14749.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e411\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eADF47159.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e526\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eAAX31279.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e428\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eO65781.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e350\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCAI79402.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e435\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e\n\u003ch3\u003eFunctional analysis\u003c/h3\u003e\n\u003cp\u003eThe function of a protein depends on its combination of domains (Fong and Marchler-Bauer \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). For functional characterization of protein the conserved domains were searched through Conserved domain database (CDD) (Marchler-Bauer et al. \u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e2011\u003c/span\u003e). The motifs were predicted using MEME suite (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://meme.nbcr.net/meme/cgi-bin/meme.cgi\u003c/span\u003e\u003cspan address=\"http://meme.nbcr.net/meme/cgi-bin/meme.cgi\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e). The parameter of motif size was set at 6 and 50 as minimum and maximum, respectively. The gene ontology classification was also done by using Uniprot database (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.uniprot.org\u003c/span\u003e\u003cspan address=\"http://www.uniprot.org\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e). Each protein was classified in three classes, biological process, molecular function and cellular component.\u003c/p\u003e\n\u003ch3\u003ePrimary Structure Analysis\u003c/h3\u003e\n\u003cp\u003ePrimary structure analysis or physico-chemical characterization was done by using the Expasy\u0026rsquo;s ProtParam server (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://web.expasy.org/protparam/\u003c/span\u003e\u003cspan address=\"https://web.expasy.org/protparam/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e). ProtParam was used to compute the biophysical and biochemical properties like isoelectric point (pI), molecular weight, instability index (Guruprasad et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e1990\u003c/span\u003e), aliphatic index (Ikai \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e1980\u003c/span\u003e) and grand average hydropathy (GRAVY) (Kyte and Doolittle \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e1982\u003c/span\u003e).\u003c/p\u003e\n\u003ch3\u003eSecondary Structure Prediction\u003c/h3\u003e\n\u003cp\u003eCFSSP (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.biogem.org/tool/chou-fasman/\u003c/span\u003e\u003cspan address=\"https://www.biogem.org/tool/chou-fasman/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) was employed for calculating the secondary structural features present in the protein sequences of galactomannan biosynthetic pathway considered in this study. The protein sequences were analyzed to predict the contribution of alpha helix, beta sheet and random coil structures (Chou and Fasman \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e1974\u003c/span\u003e) {30].\u003c/p\u003e\u003cdiv id=\"Sec8\" class=\"Section2\"\u003e\u003ch2\u003eModel building and evaluation\u003c/h2\u003e\u003cp\u003eThe three dimensional structure of the protein was predicted by threading alignment modelling program using I-TASSER server (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://zhanggroup.org/I-TASSER/\u003c/span\u003e\u003cspan address=\"https://zhanggroup.org/I-TASSER/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) (Yang et al. \u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e2015\u003c/span\u003e; Zheng et al. \u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The templates were selected from a link connecting to PDB database (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.rcsb.org/\u003c/span\u003e\u003cspan address=\"https://www.rcsb.org/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) consisting of known structures. The high resolution crystal structures of template proteins were considered for threading modelling. The template with maximum identity value and minimum e-score was selected for each protein sequence to build a model. The energy minimization was performed to stabilize the modelled protein with Swiss PDB Viewer (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://spdbv.unil.ch/\u003c/span\u003e\u003cspan address=\"https://spdbv.unil.ch/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) (Johansson et al. \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e2012\u003c/span\u003e). The model having higher C value (Confidence score) were chosen for further analysis. Structure visualization was performed through Pymol (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://pymol.org/2/\u003c/span\u003e\u003cspan address=\"https://pymol.org/2/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e).\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eStructure validation\u003c/h3\u003e\n\u003cp\u003eAfter the model generation, its quality assessment was done, based on both geometric and energetic aspects. The stereochemical properties of obtained protein model were checked using RAMPAGE server (Lovell et al. \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e2003\u003c/span\u003e). The stereochemical properties of the proteins were assessed by Ramachandran plot analysis (Ramakrishnan and Ramachandran \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e1965\u003c/span\u003e). The validation for modelled structures was done to determine whether the residues were falling in the most favoured region in the Ramachandran plot by using PROCHECK (Laskowski et al. \u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e1996\u003c/span\u003e).\u003c/p\u003e\n\u003ch3\u003eProtein ligand binding site analysis\u003c/h3\u003e\n\u003cp\u003eAll modelled proteins were analyzed for ligand binding site by using COACH server (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://zhanggroup.org/COACH/\u003c/span\u003e\u003cspan address=\"https://zhanggroup.org/COACH/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e ). This server generates complementary ligand binding sites using two comparative methods, TM-SITE and S-SITE, which recognizes ligand binding templates from BioLiP protein function database by binding specific substructure and sequence profile comparisons. These predictions will be combined with results from other methods including COFACTOR (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://zhanggroup.org/COFACTOR/\u003c/span\u003e\u003cspan address=\"https://zhanggroup.org/COFACTOR/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), FINDSITE (Brylinski and Skolnick \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e2008\u003c/span\u003e) and ConCavity (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://compbio.cs.princeton.edu/concavity/\u003c/span\u003e\u003cspan address=\"https://compbio.cs.princeton.edu/concavity/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) to generate final ligand binding site predictions.\u003c/p\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003eActive site prediction\u003c/h2\u003e\u003cp\u003eProtein-protein superposition, brings ligand into the target active site with an orientation similar to the one found in the original ligand-receptor crystal structure (Srivastava et al. \u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e2010\u003c/span\u003e). The ligand binding site of modelled protein was predicted by superimposition of known template proteins by using COACH server (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://zhanggroup.org/COACH/\u003c/span\u003e\u003cspan address=\"https://zhanggroup.org/COACH/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e ). The RMSD was calculated using the superimposition between matched pairs. Ligand showing higher RMSD value and Z-value more than 1 were used for docking study. Active site residues and ligand binding site residues are shown in Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e9\u003c/span\u003e.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 9\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eParameters used for the 3D structure, ligand binding site and active site prediction of the pathway protein by using I-TASSER server\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"17\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c13\" colnum=\"13\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c14\" colnum=\"14\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c15\" colnum=\"15\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c16\" colnum=\"16\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c17\" colnum=\"17\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eTemplate used for modeling\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eIdent 1\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eIdent 2\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eCov\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eNorm Z score\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eCluster Size\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e\u003cp\u003eC-Score\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c9\"\u003e\u003cp\u003eLig Name\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c10\"\u003e\u003cp\u003eLig binding Residues\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c11\"\u003e\u003cp\u003eC score EC\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c12\"\u003e\u003cp\u003eTM score\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c13\"\u003e\u003cp\u003eRMSDa\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c14\"\u003e\u003cp\u003eIDENa Cov\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c15\"\u003e\u003cp\u003eEC No\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c16\"\u003e\u003cp\u003eActive site res\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c17\"\u003e\u003cp\u003ePDB hit for active site\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1uasA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e0.72\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e0.63\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e0.88\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2.73\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e77\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e0.66\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003eGLA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e63, 98, 99,140, 148, 175, 177, 209, 211, 228, 232, 264\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e\u003cp\u003e0.737\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e\u003cp\u003e0.875\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e\u003cp\u003e0.62\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e\u003cp\u003e0.715\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c15\"\u003e\u003cp\u003e3.2.1.22\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c16\"\u003e\u003cp\u003e177,232\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c17\"\u003e\u003cp\u003e1uasA\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e4hg6A\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e0.18\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e0.27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e0.96\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e1.86\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e37\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e0.36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003eUDP\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e98,99,100,102,129,165,166,169,189,190,322\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e\u003cp\u003e0.203\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e\u003cp\u003e0.461\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e\u003cp\u003e5.61\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e\u003cp\u003e0.071\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c15\"\u003e\u003cp\u003e2.4.1.41\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c16\"\u003e\u003cp\u003eNA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c17\"\u003e\u003cp\u003e2d7iA\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e1pmiA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e0.39\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e0.42\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e0.96\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2.84\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e11\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e0.17\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003eKOJ\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e55,108,117,119,144,146,282,284\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e\u003cp\u003e0.627\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e\u003cp\u003e0.616\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e\u003cp\u003e2.88\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e\u003cp\u003e0.192\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c15\"\u003e\u003cp\u003e5.3.1.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c16\"\u003e\u003cp\u003e291,301\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c17\"\u003e\u003cp\u003e1qwrA\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e4ouaB\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e0.09\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e0.20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e0.93\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e1.47\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e0.14\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003eOTR\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e38,177,178,181,193,194,195,196,199\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e\u003cp\u003e0.117\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e\u003cp\u003e0.334\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e\u003cp\u003e7.77\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e\u003cp\u003e0.029\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c15\"\u003e\u003cp\u003e1.4.1.13\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c16\"\u003e\u003cp\u003eNA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c17\"\u003e\u003cp\u003e2vdcF\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003e3enkA\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e0.56\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e0.54\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e0.97\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e3.85\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e556\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e0.98\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003eNAD\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e10,12,13,14,15,34,35,36,37,38,39,63,64,65,86,87,88,90,105,128,129,130,154,158,182,183,184,185\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c11\"\u003e\u003cp\u003e0.681\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c12\"\u003e\u003cp\u003e0.908\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c13\"\u003e\u003cp\u003e1.30\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c14\"\u003e\u003cp\u003e0.424\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c15\"\u003e\u003cp\u003e5.1.3.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c16\"\u003e\u003cp\u003e14,154,183,205,238\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c17\"\u003e\u003cp\u003e2c20A\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003ctfoot\u003e\u003ctr\u003e\u003ctd colspan=\"17\"\u003e1) α-D-Galactoside galactohydrolase 2) β-1,4-Mannan Synthase 3) Phosphomannose isomerise 4) Galactomannan Galactosyltransferase 5) UDP-Galactose 4-epimerase\u003c/td\u003e\u003c/tr\u003e\u003c/tfoot\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec12\" class=\"Section2\"\u003e\u003ch2\u003eMolecular docking\u003c/h2\u003e\u003cp\u003eThe docking of ligand with modelled protein was performed by Swiss Dock (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.swissdock.ch/\u003c/span\u003e\u003cspan address=\"http://www.swissdock.ch/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) at ExPASY server. In this approach, the full flexible ligand was used for docking while keeping the protein in a fixed orientation in space. The negative and lower value of binding energy as well as more numbers of cluster involved showed favoured binding between ligand and target.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e\u003ch2\u003ePlant material and drought experiment\u003c/h2\u003e\u003cp\u003eCluster bean (\u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e) seeds were grown in pots in soil and placed in growth chamber (PGC-105 HID; Percival Scientific, Inc., 505 Research Drive, Perry, IA 5022D, USA) for the study. The temperature variation was between 25\u0026ndash;32\u0026deg;C and relative humidity was in the range of 50\u0026ndash;70%. The photosynthetic photon flux density increased gradually from 6:00 to 12:00 h which ranged from (50\u0026ndash;1300 \u0026micro;mol m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e) and then decreased till 7:00 pm to maintain light and dark cycle for the plants. The two months old plants were subjected to drought by withdrawal of water.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec14\" class=\"Section2\"\u003e\u003ch2\u003eMeasurement of relative water content\u003c/h2\u003e\u003cp\u003eTo measure relative water content of the varieties three to five mature leaves at 0, 1st, 5th, 10th day of drought and at rewatered stage were harvested early morning to measure the fresh weight. These leaf samples were kept overnight in water to measure the turgid weight (TW) the next day after they had attain turgidity and then these were kept in oven at 70\u0026deg;C for 72h. The dry weight was recorded and the relative water content RWC was calculated as\u003c/p\u003e\u003cp\u003eRWC (%) = [(FW\u0026thinsp;\u0026minus;\u0026thinsp;DW)/ (TW\u0026thinsp;\u0026minus;\u0026thinsp;DW)] \u0026times;100.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e\u003ch2\u003eMeasurement of the osmotic potential\u003c/h2\u003e\u003cp\u003eOsmotic potential of plants under different days of drought were determined in the leaves using a Wescor 5520 osmometer (Wescor, Logan, Utah, USA) as described by Ball and Oosterhuis (2005). Osmotic potential measurements were taken with leaf discs weighing approx 10-15mg each.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec16\" class=\"Section2\"\u003e\u003ch2\u003eGas exchange measurements\u003c/h2\u003e\u003cp\u003eThe net photosynthesis rate (\u003cem\u003eA\u003c/em\u003e), stomatal conductance (\u003cem\u003egs\u003c/em\u003e), transpiration (\u003cem\u003eE\u003c/em\u003e), and WUE were measured in fully expanded leaves of guar at the regular interval with portable photosynthesis system GFS-300 (Gas exchange and fluorescence measuring system). The photosynthetic photon flux density was maintained at 800 \u0026micro;mol m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e s\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, CO\u003csub\u003e2\u003c/sub\u003e level in the leaf cuvette was 400 ppm, vapour pressure deficit (VPD) level was less than 2KPa and temperature of the leaf was at 30\u0026deg;C.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec17\" class=\"Section2\"\u003e\u003ch2\u003eChlorophyll estimation\u003c/h2\u003e\u003cp\u003eThe photosynthetic pigments such as chlorophyll and carotenoid were measured using method of Wellburn(Wellburn \u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e1994\u003c/span\u003e). Leaf tissue of 20\u0026ndash;25 mg was taken each from control and drought treated plants. These were kept in 1 ml of 80% acetone in dark for pigment extraction, then OD were recorded at 470, 647 and 663 nm for chl a, chl b and carotenoids determination.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec18\" class=\"Section2\"\u003e\u003ch2\u003ePhysicochemical characterization of gum\u003c/h2\u003e\u003cp\u003eA Physiochemical property of the seed was measured as follows:\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec19\" class=\"Section2\"\u003e\u003ch2\u003eSample collection\u003c/h2\u003e\u003cp\u003eSeeds were collected from pods of guar plants of both watered and drought treated conditions. Seeds were soaked in water to remove endosperm and were kept in a refrigerator at 2\u0026ndash;4\u0026deg; C for at most 24 h prior to isolation and purification.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec20\" class=\"Section2\"\u003e\u003ch2\u003eExtraction and Isolation of the seed gum\u003c/h2\u003e\u003cp\u003eExtraction and isolation of gum from seeds of guar were done using the procedure of Liyanage et al \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2015\u003c/span\u003e (Liyanage et al. \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e2015\u003c/span\u003e). The seeds were soaked in water to remove the endosperm for gum extraction. These were then dried in oven to remove moisture .Approximately (10\u0026ndash;12 g) of the endosperm was grinded to a powder and left overnight in 50ml hexane. The powder was properly washed the next day in hexane and was filtered. The filtrate obtained was dried to remove excess hexane. 100ml water was then added to this and kept overnight. The next day it was filtered and washed in water for 2\u0026ndash;3 times. The filtrate was centrifuged at 12000rpm for 10 mins and supernatant was taken. To this supernatant add ethanol (1.5 times the supernatant). A thick substance precipitates out as gum. This precipitate was oven dried at 50\u003csup\u003e⁰\u003c/sup\u003eC till it was powder. To purify the gum, the dried gum was dissolved in isopropanol (same volume as that of gum) with continuous stirring until completely dissolved and then finally dried.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec21\" class=\"Section2\"\u003e\u003ch2\u003eDetermination of Specific Rotation of Gums\u003c/h2\u003e\u003cp\u003eEach gum sample (.01%) were filtered and equilibrated at 27\u0026deg; C of temperature. Optical rotations were determined for the three samples using polarimeter according to Dawber et al. (Dawber et al. \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e1988\u003c/span\u003e). The specific rotation of gum samples were measured in degrees.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec22\" class=\"Section2\"\u003e\u003ch2\u003eViscosity\u003c/h2\u003e\u003cp\u003eGum solutions of 0.01% were prepared by continuous stirring using a magnetic stirrer until the solution gelatinized. The actual viscosity of the gum was measured at 30\u0026deg; C at 6 rpm by a Well Brookfield Cone/Plate Microviscometer model R VT according to Stoloff et al (Stoloff \u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e1958\u003c/span\u003e). The viscometer reading in centipoise (cP) was multiplied by a factor equivalent to the spindle number that has been used.\u003c/p\u003e\u003cdiv id=\"Sec23\" class=\"Section3\"\u003e\u003ch2\u003eAsh and carbon content\u003c/h2\u003e\u003cp\u003ePercentage ash and carbon content was determined using Analyser Multi N/C (Analytic Jena, Germany) 2100S. 40\u0026ndash;50 mg of gum sample was used for estimation of carbon and ash content.\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e"},{"header":"Results and discussion","content":"\u003cdiv id=\"Sec25\" class=\"Section2\"\u003e\u003ch2\u003eFunctional analysis of pathway proteins\u003c/h2\u003e\u003cp\u003eDomains are evolutionarily conserved units in proteins and frequently correspond to recurrent structural and functional units (Basu et al. \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2008\u003c/span\u003e). The particular function of a protein depends on its combination of domains; 80% of eukaryotic proteins have more than one domain. The galactomannan biosynthesis pathway proteins were analyzed for the availability of conserved domains. The domain having least e-value was selected corresponding to each protein. The retrieved conserved domains and their position in proteins were analyzed. Galactomannan Galactosyltransferase has single domains indicating that they did not participate in the formation of multi-domain structure. However α-D-Galactoside galactohydrolase, β-1,4-Mannan Synthase, Phosphomannose isomerise, UDP-Galactose 4-epimerase has two or three domains and participated in multiple domain structure (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e2\u003c/span\u003e).Three motifs for each conserved domains were predicted. The motifs might be involved in binding site formation (Sanchita et al. 2013). The substrates of these proteins bind at these motifs of proteins (e.g enzymes). The motifs are represented in form of regular expression and web logo that explains the frequency of occurrence of residues of any motif at minimum 2 sites (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eConserved Domain found in pathway proteins\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eConserved Domain\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003ePosition in the protein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eE-value\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGH27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e56\u0026ndash;319\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e2.67e\u003csup\u003e\u0026minus;\u0026thinsp;159\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePLN02692\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e1-411\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0e\u003csup\u003e+\u0026thinsp;00\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eCESA_CaSu_A2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e91\u0026ndash;327\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e1.4e\u003csup\u003e\u0026minus;\u0026thinsp;137\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGlyco_tranf_2_3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e91\u0026ndash;327\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e1.51e\u003csup\u003e\u0026minus;\u0026thinsp;25\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePLN02288\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e14\u0026ndash;407\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0e\u003csup\u003e+\u0026thinsp;00\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eMan A\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e12\u0026ndash;404\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e5.55e\u003csup\u003e\u0026minus;\u0026thinsp;55\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eUDP_G4E_1_SDR_e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e5-334\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0e\u003csup\u003e+\u0026thinsp;00\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePLN02240\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e1-349\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0e\u003csup\u003e+\u0026thinsp;00\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePLN03181\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e1-435\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e0e\u003csup\u003e+\u0026thinsp;00\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003e\u003cb\u003eDomains with corresponding motifs and their regular expression\u003c/b\u003e\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"8\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eNo. of Motifs\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eRegular expression\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eLogo\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eE-value\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eWidth\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eSite count\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e\u003cp\u003eR e entropy\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e[RQ][YT][PP][MG]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e3.3e\u003csup\u003e+\u0026thinsp;000\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e25.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e[WR][SN][HF][SG][IC]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e9.8e\u003csup\u003e+\u0026thinsp;000\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e24.2\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e[KF][YF][SD][NM][CE]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1.2e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e24.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eA[IT][YKF]D[AL][LGD][FG][WQG][PKH]D[RHA][TFD][FT]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e4.1e\u003csup\u003e+\u0026thinsp;000\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e13\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e44.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eQ[HA]RW[SK][FC]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1.3e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e22.9\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e[SH][LS[TRID]Y[ASH][FW]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e5.3e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e20.4\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCS[VM][LK][TN]Y[KD][WQ]G\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1.5e\u003csup\u003e+\u0026thinsp;000\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e36.3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eE[IC][ID][RE]C[MI]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e3.8e\u003csup\u003e+\u0026thinsp;000\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e25.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e[SI]Q[FA][HPD]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e3.7e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e23.9\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eI[GED]E[MID][CP]R[DG][IQ][YWP][NR][WNA]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e8.4e\u003csup\u003e\u0026minus;\u0026thinsp;002\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e11\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e39.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eE[QL][NE][WK][IA]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e6.7e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e23.8\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e[QP][QL]V[MA][VA][GRRP]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e8.1e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e34.2\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eI[TL][LH][VF]TG[SC][QP][KC][RN][GC]H\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e7.2e\u003csup\u003e\u0026minus;\u0026thinsp;002\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e13\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e51.9\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eD[VC]W[VN][SG]M\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e1.6e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e24.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eH[SG]W[TE][GL]\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e6.0e\u003csup\u003e+\u0026thinsp;001\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c8\"\u003e\u003cp\u003e23.8\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cdiv id=\"Sec26\" class=\"Section3\"\u003e\u003ch2\u003eGene ontology classification of pathway proteins\u003c/h2\u003e\u003cp\u003eThe gene ontological classification of the proteins was analyzed in terms of biological process, molecular function and cellular component (Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e4\u003c/span\u003e). α-D-Galactoside galactohydrolase, Phosphomannose isomerise, UDP-Galactose 4-epimerase, enzymes involved in the carbohydrate metabolism while β-1,4-Mannan Synthase, Galactomannan Galactosyltransferase are the membrane bound enzymes and participated in cell wall organisation, biogenesis and degradation process. Phosphomannose isomerise, mannan synthase participated in mannan biosynthesis but PMI also involved in Zn ion binding. UDP-Galactose 4-epimerase catalyzes the formation of UDP galactose from UDP-glucose or fructose.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003e\u003cb\u003eGene ontology classification of enzymes is dividing into three group biological process, molecular function and cellular component\u003c/b\u003e.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eProtein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eBiological Process\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eMolecular Function\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eCellular Component\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCarbohydrate metabolic process, catalytic and enzymatic activity, α galactosidase activity and raffinose α galactosidase activity\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eGlycosidase, Hydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCell wall biogenesis and degradation, metabolic process and cell wall organization, GDP-mannose + (mannan)(n)\u0026thinsp;=\u0026thinsp;GDP + (mannan)(n\u0026thinsp;+\u0026thinsp;1).\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eTransferase activity, transferring glycosyl groups\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003eGolgi apparatus, and integral component of membrane\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCarbohydrate metabolic process, GDP mannose biosynthetic process\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eMannose-6-phosphate isomerase activity, zinc ion binding\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCarbohydrate metabolic process, galactose metabolic process, Ligand NAD\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eCoenzyme binding, UDP-glucose 4-epimerase activity, Isomerase activity\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003eMetabolic process, cell wall organisation\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003eTransferase activity, transferring hexosyl groups\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003eGolgi apparatus, and integral component of membrane\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec27\" class=\"Section3\"\u003e\u003ch2\u003ePhysicochemical properties of proteins\u003c/h2\u003e\u003cp\u003eParameters computed using Expasy\u0026rsquo;s ProtParam tool given in Table\u0026nbsp;\u003cspan refid=\"Tab6\" class=\"InternalRef\"\u003e5\u003c/span\u003e. The calculated isoelectric point (pI) is useful because at pI, solubility is least and mobility in an electro focusing system is zero. Isoelectric point (pI) is the pH at which the surface of protein is covered with charge but net charge of protein is zero. At pI proteins are stable and compact. The computed pI value of all protein except α-D-Galactoside galactohydrolase is more than 7 indicating the net electrical charge of proteins is zero at basic pH value. However, α-D-Galactoside galactohydrolase showed less than 7 (pI\u0026thinsp;\u0026lt;\u0026thinsp;7) reveals that net electrical charge is zero at acidic pH value. The computed isoelectric point (pI) will also be useful for developing buffer system for purification by isoelectric focusing method. The instability index provides an estimate of the stability of protein in a test tube. This method assigns a weight value of instability. Using these weight values, it is possible to compute an instability index. A protein whose instability index is smaller than 40 is predicted as stable, a value above 40 predicts that the protein may be unstable (Guruprasad et al. \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e1990\u003c/span\u003e). The instability index value for these proteins was found to be ranging from 28.57 to 40.36. β-1, 4-Mannan Synthase and Phosphomannose isomerise showed comparatively high instability index 40. The very high aliphatic index of protein sequences indicates that β-1, 4-Mannan Synthase and Phosphomannose isomerise proteins may be stable for a wide temperature range. The lower thermal stability of α-D-Galactoside galactohydrolase, UDP-Galactose 4-epimerase and Galactomannan Galactosyltransferase was indicative of a more flexible structure when compared to other protein. The Grand Average Hydropathy (GRAVY) value for a protein is calculated as the sum of hydropathy values of all amino acids, divided by the number of residues in the sequence. GRAVY indices were found in the range from \u0026minus;\u0026thinsp;0.592 to 0.106. This low range of value indicates the possibility of better interaction with water. Negative value of hydropathy showed hydrophilic proteins while positive showed hydrophobic nature of the protein (Kyte and Doolittle \u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e1982\u003c/span\u003e). Except β-1, 4-Mannan synthase all studied proteins showed negative value of hydropathy and these are hydrophilic nature towards water.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab6\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003e\u003cb\u003ePhysicochemical parameter of pathway proteins is computed by using Expasy\u0026rsquo;s ProtParam tool.\u003c/b\u003e\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAA\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eMol wt\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003epI\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eInstability Index\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eAliphatic Index\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eGRAVY\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e411\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e45135.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e4.97\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e32.11\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e76.69\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e-0.359\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e526\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e60677.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e9.28\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e40.35\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e100.78\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e0.106\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e428\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e47773.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e6.60\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e40.36\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e89.72\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e-0.223\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e350\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e38373.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e6.72\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e28.57\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e87.74\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e-0.198\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e435\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e50956.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e7.98\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e35.57\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e72.44\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c7\"\u003e\u003cp\u003e-0.592\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e\u003cdiv id=\"Sec28\" class=\"Section2\"\u003e\u003ch2\u003eStructural analysis of modelled pathway proteins\u003c/h2\u003e\u003cp\u003eThe secondary structure of proteins was analyzed for considering relative availability of alpha helix, extended strand and random coils. The percentage distribution of predicted secondary structure features are represented in Table\u0026nbsp;\u003cspan refid=\"Tab7\" class=\"InternalRef\"\u003e6\u003c/span\u003e. The results revealed that the predicted alpha helix dominated among secondary structure elements followed by extended strand, random coils for all protein sequences. Present study findings were similar to the earlier results reported in \u003cem\u003eWithania somnifera\u003c/em\u003e (Sanchita et al. 2014).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab7\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 6\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eSecondary structure elements identified in proteins.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"4\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eAlpha Helix (%)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eExtended strand (%)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eRandom Coil (%)\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e62.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e59.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e14.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e85.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e53.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e9.5\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e82.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e67.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e13.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e68.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e38.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e12.9\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e62.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e61.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e13.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003eThree-dimensional structures were predicted for proteins where such data is unavailable. There is lack of experimental structures for the proteins considered. The modelling of the three dimensional structure of these proteins were performed by threading alignment web server, I-TASSER. The template having maximum identity (maximum alignment coverage) and highest c value was selected and models were created using I-TASSER server. The model having lowest energy was considered for further study (Fig.\u0026nbsp;2). The stereochemical quality and accuracy of the predicted models were evaluated using Ramachandran map calculations computed with the PROCHECK program. The result revealed that the modelled structure for all protein sequences have 76.5%, 72.7%, 74.3%, 76.5%, 80.9% and 56.2% residues for α-D-Galactoside galactohydrolase, β-1,4-Mannan Synthase, Phosphomannose isomerise, UDP-Galactose 4-epimerase and Galactomannan Galactosyltransferase respectively in most favoured region (Table\u0026nbsp;\u003cspan refid=\"Tab8\" class=\"InternalRef\"\u003e7\u003c/span\u003e). The identified Ramachandran plots represent a good quality of predicted models. The structures were naturally stable with low energy value. The RMSD values identified through superimposition between selected template structure and modelled structure.\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab8\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 7\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003ePercentage distribution of amino acid residues in ramachandran plot.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"5\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eProtein\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eResidues in most favored region (%)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eResidues in additionally allowed region (%)\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eResidues in generously allowed region\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eResidues in disallowed region\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e76.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e19.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e3.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1.4\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e72.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e19.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e4.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e3.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e74.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e21.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e2.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e1.9\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e80.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e15.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e3.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e0.3\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e56.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e32.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e7.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e3.6\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec29\" class=\"Section2\"\u003e\u003ch2\u003eLigand binding site prediction for modelled proteins\u003c/h2\u003e\u003cp\u003eModelled proteins of galactomannan biosynthetic pathway were analyzed for ligand binding site prediction. PMI showed minimum number of ligand binding residues while UDP-G4E has maximum. α-D-Galactoside, MS, PMI, GT, UDP-G4E have shown GLA, UDP, Zn, OTR and NAD as probable ligands, respectively (Fig.\u0026nbsp;3). All modelled proteins showed more than 88% sequence coverage of templates. GT have minimum clusters size while UDP-G4E showed maximum templates in a cluster (Table\u0026nbsp;\u003cspan refid=\"Tab9\" class=\"InternalRef\"\u003e8\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab9\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 8\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eTrans-membrane activity of pathway proteins\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"7\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eNo of transmembrane helices\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eStart from\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eEnd to\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eHelices length\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eScore\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eOrientation\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eα-D-Galactoside galactohydrolase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e124\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e143\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2255\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eo-i\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eβ-1,4-Mannan Synthase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e78\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e100\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e1901\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eo-i\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e409\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e428\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e25554\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003ei-o\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e438\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e467\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e30\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e771\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eo-i\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e548\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e571\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2160\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003ei-o\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e303\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e322\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e832\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eo-i\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e413\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e432\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e630\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003ei-o\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e50\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e69\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e20\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e516\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003ei-o\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e112\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e133\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e22\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e752\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eo-i\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e23\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e43\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e21\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e527\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003ei-o\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e\u003cp\u003e77\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e\u003cp\u003e98\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e\u003cp\u003e22\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\".\" colname=\"c6\"\u003e\u003cp\u003e2531\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003ei-o\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eActive site prediction in the modelled proteins\u003c/h3\u003e\n\u003cp\u003eAll modelled proteins were also analyzed for active sites (Fig.\u0026nbsp;4). MS, PMI and GT did not show any active site residue while α-D-Galactoside and UDP-G4E have 2 and 5 active site residues, respectively (Table\u0026nbsp;\u003cspan refid=\"Tab9\" class=\"InternalRef\"\u003e8\u003c/span\u003e). MS, PMI and GT also showed less c value for the active site prediction. These are membrane proteins and for membrane proteins c value is always less. C\u003csub\u003eScore\u003c/sub\u003e\u003csup\u003eEC\u003c/sup\u003e is the confidence score for the EC number prediction. C\u003csub\u003eScore\u003c/sub\u003e\u003csup\u003eEC\u003c/sup\u003e values range between 0\u0026ndash;1, where a higher score indicates a more reliable EC number prediction. TM score is a measure of global structure similarity between query and template protein. RMSD\u003csup\u003ea\u003c/sup\u003e is the RMSD between residues that are structurally aligned by TM align. IDEN\u003csup\u003ea\u003c/sup\u003e is the percentage sequence identity in the structurally aligned region.\u003c/p\u003e\u003cdiv id=\"Sec31\" class=\"Section2\"\u003e\u003ch2\u003eDocking study of pathway protein with their ligands\u003c/h2\u003e\u003cp\u003eMolecular docking can fit molecules together with a favorable configuration to form a complex system. The structural information from the theoretical modeled complex can help us to understand the catalytic mechanism of an enzyme. The modelled pathway proteins were studied for docking with their probable ligand respectively. UDP-G4E showed minimum enthalpy energy and fitness energy thus it forms more stable UDP galactose which further converted in to galactomannans (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e9\u003c/span\u003e). However, PMI showed comparatively higher enthalpy change and catalyze reversible reaction and forms less stable compound. Ligand binding at appropriate configuration (with lowest enthalpy change) is given in Fig.\u0026nbsp;5.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec32\" class=\"Section2\"\u003e\u003ch2\u003ePhysiological performance of the guar plants under drought stress\u003c/h2\u003e\u003cp\u003eMilan variety of guar was imposed to drought for 10 days. Relative water content of the leaf dropped up to 50% at the 10th day of drought (DD) (Fig.\u0026nbsp;6A). Osmotic potent also decreased under the drought and plants were not fully recovered after the one week of recovery (Fig.\u0026nbsp;7B). The photosynthesis rate (\u003cem\u003eA\u003c/em\u003e) was decreased by 66.2% in 10DD, however stomatal conductance (\u003cem\u003eg\u003c/em\u003e\u003csub\u003e\u003cem\u003es\u003c/em\u003e\u003c/sub\u003e) and transpiration rate (\u003cem\u003eE\u003c/em\u003e) did not change significantly under drought stress (Fig.\u0026nbsp;7A, B, C). Water use efficiency (\u003cem\u003eWUE\u003c/em\u003e) decreased by 61% under drought stress. However, fully recovered in one of recovery (Fig.\u0026nbsp;7D). The chlorophyll content decreased up to 60% in comparison to watered plants. Chlorophyll a/b ratio did not changed significantly under drought stress (Fig.\u0026nbsp;8A, B). carotenoid content increased dramatically at the 5th DD and did not recovered after the recovery period (Fig.\u0026nbsp;8C). Anthocyanin content increased upto 80 \u0026micro;molper gram of fresh weight. Further it decreased after the rewatering of one week (Fig.\u0026nbsp;8D). Drought causes decrease in water content which is accompanied by loss of turgor and wilting, closure of stomata, reduction in photosynthesis, and interference with many other basic metabolic processes (Singh et al. \u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). Drought stress primarily decreases the photosynthesis rate due to decrease in internal CO\u003csub\u003e2\u003c/sub\u003e concentration resulting from stomatal closure (Chaves et al. \u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e2009\u003c/span\u003e; Vishnyakova et al. \u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Chlorophyll and carotenoid are light harvesting photosynthetic pigments which showed changes to avoid the inhibition in photosynthesis reaction centers under drought stress. These pigments also degrade which leads to deterioration of thyllakoid membranes (Munn\u0026eacute;-Bosch and Alegre \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e2000\u003c/span\u003e). Carotenoids act as accessory light harvesting pigments thus transferring light energy to chlorophylls. Anthocyanins are another class of pigments which also have photoprotective effect. Under stress conditions to avoid photooxidative damage to plant tissues anthocyanins functions as antioxidants by scavenging reactive oxygen radicals (Ranjan et al. \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). These act as light attenuators by not transferring the absorbed light energy to the photosynthetic machinery of the plants thus protecting underlying cells from high irradiance (Hogewoning et al. \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e2012\u003c/span\u003e; Zhu et al. \u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e2016\u003c/span\u003e). Our study revealed that the anthocyanin content is found to increase under drought which is a protective mechanism of plant against stress.\u003c/p\u003e\u003cdiv id=\"Sec33\" class=\"Section3\"\u003e\u003ch2\u003eGene expression of the proteins involved in the galactomannan biosynthesis\u003c/h2\u003e\u003cp\u003eFru-6-P converts into Man-6-P by phosphomannose isomerise (PMI) which also functions in the reverse direction for the utilization of mannose. Expression of PMI is decreased by 15.8 fold in comparison to watered plants (Fig.\u0026nbsp;9). UDP-G4E is the enzyme which forms the direct precursor for galactomannan biosynthesis, UDP-D-galactose. In this study expression value was upregulated by 5.8 fold in comparison to control plants (Fig.\u0026nbsp;9). UDP-galactose-dependent GT transfers a galactose residue through a α-linkage to a mannose near the nonreducing end of the growing mannan chain. Expression of GT is upregulated by 20 fold while MS increased by 60.5 fold in comparison to watered plans (Fig.\u0026nbsp;9). Galactosidase is a enzyme which hydrolyse the galactose residue of galactomannan. Expression of galactosidase is down regulated by -30.6 fold in comparison to control plants (Fig.\u0026nbsp;9). Overall enzymes involved in the galactomannan biosynthesis are upregulated. However, enzymes associated with hydrolysis are downregulated. Formation of seed and fruit influenced by various metabolic processes occurred in leaves and translocated in to seeds. Drought stress affect the physiological and biochemical process which involved in the grain filling and grain quality (Sehgal et al. \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e2018\u003c/span\u003e; Boote et al. \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e2023\u003c/span\u003e). Although seed yield were decreased under water stress but seed gum yield increased under drought stress.\u003c/p\u003e\u003c/div\u003e\u003cdiv id=\"Sec34\" class=\"Section3\"\u003e\u003ch2\u003eEffect of drought on gum yield and its physiochemical properties\u003c/h2\u003e\u003cp\u003eThe physiochemical properties of seed gum were studied. The yield of gum was increased with 1.9 fold under drought stress (Table\u0026nbsp;\u003cspan refid=\"Tab10\" class=\"InternalRef\"\u003e11\u003c/span\u003e). The specific rotation of .01% sample was decreased by 1.1 fold under drought stress. Drought stress increased relative conductivity of gum samples. Percentage of ash in gum is believed to be the measure of mineral quantities in the guar seed (Dakia et al. \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e2008\u003c/span\u003e; Zammel et al. \u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e2021\u003c/span\u003e). The percentage of ash showed significant decrease by 2.2 fold under water stress. Measurements of viscosity showed increased by 1.2 fold (Table\u0026nbsp;\u003cspan refid=\"Tab10\" class=\"InternalRef\"\u003e11\u003c/span\u003e). The specific rotation of organic molecules such as carbohydrates is related to their structure and is a characteristic property of the substance (Sathyanarayana and Stevens \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e1987\u003c/span\u003e). The specific rotation of gum is thus considered as the most important criterion of purity and qualitative analysis. The percentage ash content indicates the presence of inorganic elements and mineral quantities in gum. The viscosity of any liquid represents its resistance to shearing and stirring. It is an important analytical parameter which depends on shape and size of the macromolecule (Anderson \u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1978\u003c/span\u003e). Viscosity is the most characteristic feature of guar gum it is due to its property to hydrate in aqueous solutions. Viscosity of the gum depends on several factors such as temperature, time, pH, concentration and ionic strength and type of agitation (Mudgil et al. \u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e2014\u003c/span\u003e). It was observed that drought increased viscosity of the gum solutions. This property of viscosity is due to galactomannan which may increase during drought stress (Hoppe and Goswami \u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e1999\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab10\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 11\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eGum yield and rheological properties of seed gum\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"3\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"char\" char=\"\u0026plusmn;\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eParameter\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eControl\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eDrought\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eGum Yield (%)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e\u003cp\u003e8.67\u0026thinsp;\u0026plusmn;\u0026thinsp;1.53\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e\u003cp\u003e15.87\u0026thinsp;\u0026plusmn;\u0026thinsp;1.87\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eOptical Rotation (\u0026micro;S cm\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e\u003cp\u003e60.51\u0026thinsp;\u0026plusmn;\u0026thinsp;3.89\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e\u003cp\u003e54.78\u0026thinsp;\u0026plusmn;\u0026thinsp;4.23\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eViscosity (cP)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e\u003cp\u003e25.89\u0026thinsp;\u0026plusmn;\u0026thinsp;3.78\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e\u003cp\u003e32.45\u0026thinsp;\u0026plusmn;\u0026thinsp;2.67\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u003cp\u003eCarbon content (%)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c2\"\u003e\u003cp\u003e34.75\u0026thinsp;\u0026plusmn;\u0026thinsp;2.35\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"char\" char=\"\u0026plusmn;\" colname=\"c3\"\u003e\u003cp\u003e39.56\u0026thinsp;\u0026plusmn;\u0026thinsp;33.12\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\u003c/div\u003e"},{"header":"Conclusion","content":"\u003cp\u003e\u003cem\u003eCyamopsis tetragonoloba\u003c/em\u003e is an important legume crop used for medicine and seed gum. Seed gum is made up of galactomannan which is a polysaccharide. It is synthesized by a divert pathway from the central carbohydrate metabolism. Drought stress affects physiological and biochemical metabolism. It diverted carbon metabolism towards sink tissue. Relative water content, photosynthesis rate, transpiration rate, chlorophyll content decreased under water stress. However carotenoid and anthocyanin content increased during drought. These act as light attenuators by not transferring the absorbed light energy to the photosynthetic machinery of the plants thus protecting underlying cells from high irradiance. \u0026alpha;-D-Galactoside galactohydrolase, \u0026beta;-1,4-Mannan Synthase (MS), Phosphomannose isomerise (PMI), UDP-Galactose 4-epimerase (UDP-G4E) and Galactomannan Galactosyltransferase (GGT) are the proteins involved in galactomannan biosynthesis. Therefore these proteins are studied in silico and gene expression analysis. Three-dimensional structures, ligand binding site, active sites were also predicted for these pathway enzymes. The physico-chemical properties such as pI, AI, GRAVY, instability index, hydrophobicity and transmembrane activity were also studied. The current information will provide a platform to know the structural and functional attributes of these proteins for further studies.\u003c/p\u003e\n\u003cdiv class=\"gridtable\"\u003e\n \u003ctable id=\"Tab11\" border=\"1\" class=\"fr-table-selection-hover\"\u003e\n \u003ccaption language=\"En\"\u003e\n \u003cdiv class=\"CaptionNumber\"\u003eTable 10\u003c/div\u003e\n \u003cdiv class=\"CaptionContent\"\u003e\n \u003cp\u003eDocking parameters for pathway proteins\u003c/p\u003e\n \u003c/div\u003e\n \u003c/caption\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eName of protein\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eCluster\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eElement\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eFull fitness (Kcal/mol)\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eEstimated ∆G (Kcal/mol)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026alpha;-D-Galactoside galactohydrolase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e37\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-1231.78\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-6.06\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u0026beta;-1,4-Mannan Synthase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e32\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-936.42\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-10.25\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003ePhosphomannose isomerase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e0\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-1689.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-5.23\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eUDP-Galactose 4-epimerase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-1929.91\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-11.17\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003eGalactomannan Galactosyltransferase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-1212.81\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"char\"\u003e\n \u003cp\u003e-6.19\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003ch2\u003eConflict of interest\u003c/h2\u003e\u003cp\u003eThe author declares that no conflicting interests exist.\u003c/p\u003e\u003c/p\u003e\u003ch2\u003eFunding:\u003c/h2\u003e\u003cp\u003eResearch in CSIR-CIMAP laboratory was supported by HRDG CSIR New Delhi, India (Grant No. Nehru PDF/LS/ EMR I/04/2017). RS is grateful to CSIR, New Delhi, India for Nehru Postdoctoral Fellowship. Author would like to thanks the Director CSIR-NBRI for allowing taking all the physiological measurements in Plant physiology lab of CSIR-NBRI.\u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eRS designed and conceptualized the research, performed the identification, analysis and physiological study of the control and drought treated plants. She also performed docking between protein and substrate and critically revised manuscript. All authors read and approved the manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eAnderson DMW (1978) Chemotaxonomic Aspects of the Chemistry of Acacia Gum Exudates. Kew Bull 32:529. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.2307/4109654\u003c/span\u003e\u003cspan address=\"10.2307/4109654\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBasu MK, Poliakov E, Rogozin IB (2008) Domain mobility in proteins: functional and evolutionary implications. Brief Bioinform 10:205\u0026ndash;216. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1093/bib/bbn057\u003c/span\u003e\u003cspan address=\"10.1093/bib/bbn057\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBoote KJ, Hoogenboom G, Ale S et al (2023) Adapting the CROPGRO model to simulate growth and yield of guar, Cyamopsis tetragonoloba L, an industrial legume crop. 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Tree Physiol 36:1092\u0026ndash;1104. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1093/treephys/tpw047\u003c/span\u003e\u003cspan address=\"10.1093/treephys/tpw047\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-7730051/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7730051/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eCyamopsis tetragonoloba (guar or cluster bean) is an important arid legume valued for its medicinal properties and industrial gum production. The principal constituent of guar gum is galactomannan, a polysaccharide composed of galactose and mannose, synthesized in the seed endosperm through the galactomannan biosynthetic pathway. Despite its economic significance, little information is available on the structural and functional aspects of enzymes involved in this pathway. In the present study, key enzymes including α-D-galactoside galactohydrolase, β-1,4-mannan synthase (MS), phosphomannose isomerase (PMI), UDP-galactose 4-epimerase (UDP-G4E), and galactomannan galactosyltransferase (GGT) were characterized using bioinformatic approaches. Secondary and three-dimensional structures were predicted along with ligand-binding and active site identification. Physicochemical properties such as isoelectric point (pI), aliphatic index (AI), grand average of hydropathicity (GRAVY), instability index, hydrophobicity, and transmembrane activity were also analyzed, providing valuable insights into enzyme stability and functionality.\u003c/p\u003e\u003cp\u003eTo complement the computational analysis, guar plants were subjected to drought stress to examine physiological and gum-related responses. Relative water content, photosynthetic rate, and transpiration rate significantly decreased under water deficit, whereas anthocyanin content increased, suggesting enhanced antioxidant defense against reactive oxygen species (ROS). Pod yield was reduced under stress; however, seed gum yield and viscosity increased. Furthermore, expression of genes involved in galactomannan biosynthesis was upregulated under drought conditions, indicating a stress-induced shift towards enhanced gum accumulation.\u003c/p\u003e\u003cp\u003eOverall, this multidisciplinary study integrates computational enzyme characterization with physiological and biochemical analyses, providing a comprehensive understanding of galactomannan biosynthesis and drought-induced gum modulation in C. tetragonoloba. These findings may facilitate targeted crop improvement strategies and industrial applications of guar gum under climate-stress scenarios.\u003c/p\u003e","manuscriptTitle":"Integrating computational and physiological analyses of galactomannan biosynthesis in Cyamopsis tetragonoloba","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-10-16 20:07:30","doi":"10.21203/rs.3.rs-7730051/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"1a4d8ac7-d12e-4505-a1c1-1dfeeb598dab","owner":[],"postedDate":"October 16th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2026-03-19T13:56:51+00:00","versionOfRecord":[],"versionCreatedAt":"2025-10-16 20:07:30","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7730051","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7730051","identity":"rs-7730051","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}