Functional Characterization of a Thermostable Amylosucrase from Bifidobacterium boum for Efficient Turanose...

Amylosucrase efficiently synthesizes sucrose isomers, including turanose, a low-glycemic functional sweetener. In this study, an amylosucrase from Bifidobacterium boum (BbAS) was identified through sequence-based analysis, which predicted favorable thermostability. Molecular dynamics simulations revealed that the B'-domain residues at the active-site entrance exhibited reduced flexibility in the turanose-bound state compared to the apo form, suggesting that turanose-induced stabilization of the active-site entrance contributes to the enhanced isomerization efficiency of BbAS. Experimental characterization confirmed these computational predictions by demonstrating optimal activity at 50 °C, while maintaining high residual activity during prolonged incubation at 45 °C. Kinetic analysis of BbAS with sucrose further revealed a biphasic non-Michaelian pattern, with a 3-fold increase in kcat,app above 49.1 mM. Notably, fructose supplementation shifted catalysis toward isomerization, achieving a turanose conversion rate of 61.3% while suppressing α-glucan formation to below 3.6%. These findings establish BbAS as a thermostable and industrially promising biocatalyst for efficient turanose production.