THA_1941 from Thermosipho africanus : A Thermostable β-1,3-Glucan Phosphorylase for Efficient β-1,3-Glucan Synthesis

Abstract β-1,3-Glucan phosphorylases capable of utilizing glucose as a priming substrate are key biocatalysts for the synthesis of functional β-1,3-glucan. In this study, we identified THA_1941 from Thermosipho africanus (TaβGP) as a GH161 β-1,3-glucan phosphorylase exhibiting robust synthetic activity towards glucose, as confirmed by 13C nuclear magnetic resonance, liquid chromatography-mass spectrometry, and sequence and structural analyses. TaβGP displayed exceptional thermostability, retaining 93% of its activity at 60 °C for 180 h, and showed broad pH tolerance ranging from pH 5.0 to 10.0, surpassing the performance of previously reported homologs. In addition, TaβGP exhibited broad substrate flexibility, accepting both α- and β-linked disaccharides, and demonstrated strong resistance to metal ions and lignocellulose-derived inhibitors. In the presence of 150 mM glucose 1-phosphate as the donor substrate, TaβGP synthesized β-1,3-glucan with a tunable average degree of polymerization (10–32), depending on the concentration of glucose used as the primer. The combination of thermostability, inhibitor resistance, and substrate versatility makes TaβGP a promising biocatalyst for the economically viable and environmentally sustainable synthesis of β-1,3-glucan from non-food biomass sources. Competing Interest Statement The authors have declared no competing interest.