A synergistic enzymatic strategy of papain and transglutaminase to engineer the structure and texture of high-moisture plant-based meat analogs

A synergistic enzymatic strategy of papain and transglutaminase to engineer the structure and texture of high-moisture plant-based meat analogs | 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 A synergistic enzymatic strategy of papain and transglutaminase to engineer the structure and texture of high-moisture plant-based meat analogs Ning Wang, Shanshan Liu, Jie Yu, Xiaoxuan Li, Fuhui Qi, Kaiyue Yuan, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-9037636/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 10 You are reading this latest preprint version Abstract This study developed a novel two-step enzymatic modification strategy involving papain hydrolysis followed by microbial transglutaminase (TGase) cross-linking to enhance the functionality of soy protein isolate (SPI) for high-moisture meat analogue (HMMA) production. By systematically optimizing the degree of hydrolysis (DH) and cross-linking conditions, it was found that moderate hydrolysis (DH = 3.56%) combined with TGase cross-linking (30 U/g) significantly improved the hydration and emulsifying properties of SPI. High-moisture extrusion of the modified SPI resulted in HMMA with a more ordered and anisotropic fibrous structure, as evidenced by a fibrous degree up to 1.86, an anisotropy index up to 1.68, enhanced hardness and springiness by 49.50% and 26.90%. These enhancements can be attributed to the rearrangement and covalent cross-linking of protein molecules, as well as the increased content of ordered structures such as β -folds and S-S bonds and improved water immobilization. Furthermore, the in vitro protein digestibility increased from 81.80% to 92.00% (enzymatic hydrolyzed groups) and 87.10% (enzymatic hydrolysis-cross-linking groups), indicating superior nutritional quality. These findings demonstrate that the synergistic "enzymatic hydrolysis-crosslinking" strategy effectively restructured SPI, providing a promising approach for producing plant-based meat alternatives with meat-like texture and enhanced performance. soy protein isolate high-moisture extrusion enzymatic modification plant-based meat analogs fibrous structure Full Text Additional Declarations No competing interests reported. Supplementary Files GraphicalAbstract.tif Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 03 Apr, 2026 Reviews received at journal 03 Apr, 2026 Reviews received at journal 31 Mar, 2026 Reviewers agreed at journal 22 Mar, 2026 Reviewers agreed at journal 20 Mar, 2026 Reviewers agreed at journal 17 Mar, 2026 Reviewers invited by journal 17 Mar, 2026 Editor assigned by journal 12 Mar, 2026 Submission checks completed at journal 11 Mar, 2026 First submitted to journal 05 Mar, 2026 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. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-9037636","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":607946474,"identity":"afbde1fa-9225-4811-97c8-457bdda75fbd","order_by":0,"name":"Ning Wang","email":"","orcid":"","institution":"Tianjin University of Science and Technology","correspondingAuthor":false,"prefix":"","firstName":"Ning","middleName":"","lastName":"Wang","suffix":""},{"id":607946475,"identity":"6d7a04ab-cdbc-41a8-b94a-efa5fe3feb91","order_by":1,"name":"Shanshan Liu","email":"","orcid":"","institution":"Tianjin University of Science and 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By systematically optimizing the degree of hydrolysis (DH) and cross-linking conditions, it was found that moderate hydrolysis (DH\u0026thinsp;=\u0026thinsp;3.56%) combined with TGase cross-linking (30 U/g) significantly improved the hydration and emulsifying properties of SPI. High-moisture extrusion of the modified SPI resulted in HMMA with a more ordered and anisotropic fibrous structure, as evidenced by a fibrous degree up to 1.86, an anisotropy index up to 1.68, enhanced hardness and springiness by 49.50% and 26.90%. These enhancements can be attributed to the rearrangement and covalent cross-linking of protein molecules, as well as the increased content of ordered structures such as \u003cem\u003eβ\u003c/em\u003e-folds and S-S bonds and improved water immobilization. Furthermore, the \u003cem\u003ein vitro\u003c/em\u003e protein digestibility increased from 81.80% to 92.00% (enzymatic hydrolyzed groups) and 87.10% (enzymatic hydrolysis-cross-linking groups), indicating superior nutritional quality. 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