School of Life Sciences, Institute of Aquatic Economic Animals and Guangdong Provincial Key Laboratory for Aquatic Economic Animals, National Engineering Center for Marine Biotechnology of South China Sea, China. Electronic address: [Email]
β-Glucosidase is the rate-limiting component of a cellulase-hydrolyzing reaction. Thermostability and glucose-tolerance are two critical criteria of the enzyme, which practically determine its performance in industrial applications. In this study, a thermostable and glucose-tolerant β-glucosidase (named Bgl1317) belonging to the glycoside hydrolase family 1 was acquired from a metagenomic library of Turpan soil through functional screening. Bgl1317 showed excellent thermostability and glucose-tolerance and its crystal structure was subsequently determined at a high resolution. Rational design based on the structure was conducted, producing three beneficial mutations A397R, L188A and A262S. While A397R improved the cellobiose activity by 80%, L188A and A262S increased the IC50 value of glucose from 0.8 to 1.5 M. The residues that may play a role in glucose-tolerance of GH1 β-glucosidases were summarized and the performances of glucose-tolerant β-glucosidases reported in recent years were discussed and compared. This study provides insights into enzymatic properties of Bgl1317 for engineering it into a powerful catalyst and β-glucosidases in general.
