Key roles of β-glucosidase BglA for the catabolism of both laminaribiose and cellobiose in the lignocellulolytic bacterium Clostridium thermocellum

Yan Xiao; Sheng Dong; Ya-Jun Liu; Chun You; Yingang Feng; Qiu Cui

doi:10.1016/j.ijbiomac.2023.126226

Key roles of β-glucosidase BglA for the catabolism of both laminaribiose and cellobiose in the lignocellulolytic bacterium Clostridium thermocellum

Int J Biol Macromol. 2023 Oct 1:250:126226. doi: 10.1016/j.ijbiomac.2023.126226. Epub 2023 Aug 7.

Authors

Yan Xiao¹, Sheng Dong¹, Ya-Jun Liu¹, Chun You², Yingang Feng³, Qiu Cui⁴

Affiliations

¹ CAS Key Laboratory of Biofuels, Shandong Provincial Key Laboratory of Synthetic Biology, Shandong Engineering Laboratory of Single Cell Oil, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, China; Shandong Energy Institute, Qingdao, China; Qingdao New Energy Shandong Laboratory, Qingdao, China; Dalian National Laboratory for Clean Energy, Qingdao, China; University of Chinese Academy of Sciences, Beijing, China.
² Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, China.
³ CAS Key Laboratory of Biofuels, Shandong Provincial Key Laboratory of Synthetic Biology, Shandong Engineering Laboratory of Single Cell Oil, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, China; Shandong Energy Institute, Qingdao, China; Qingdao New Energy Shandong Laboratory, Qingdao, China; Dalian National Laboratory for Clean Energy, Qingdao, China; University of Chinese Academy of Sciences, Beijing, China. Electronic address: fengyg@qibebt.ac.cn.
⁴ CAS Key Laboratory of Biofuels, Shandong Provincial Key Laboratory of Synthetic Biology, Shandong Engineering Laboratory of Single Cell Oil, Qingdao Institute of Bioenergy and Bioprocess Technology, Chinese Academy of Sciences, Qingdao, China; Shandong Energy Institute, Qingdao, China; Qingdao New Energy Shandong Laboratory, Qingdao, China; Dalian National Laboratory for Clean Energy, Qingdao, China; University of Chinese Academy of Sciences, Beijing, China. Electronic address: cuiqiu@qibebt.ac.cn.

PMID: 37558019
DOI: 10.1016/j.ijbiomac.2023.126226

Abstract

The thermophilic bacterium Clostridium thermocellum efficiently degrades polysaccharides into oligosaccharides. The metabolism of β-1,4-linked cello-oligosaccharides is initiated by three enzymes, i.e., the cellodextrin phosphorylase (Cdp), the cellobiose phosphorylase (Cbp), and the β-glucosidase A (BglA), in C. thermocellum. In comparison, how the oligosaccharides containing other kinds of linkage are utilized is rarely understood. In this study, we found that BglA could hydrolyze the β-1,3-disaccharide laminaribiose with much higher activity than that against the β-1,4-disaccharide cellobiose. The structural basis of the substrate specificity was analyzed by crystal structure determination and molecular docking. Genetic deletions of BglA and Cbp, respectively, and enzymatic analysis of cell extracts demonstrated that BglA is the key enzyme responsible for laminaribiose metabolism. Furthermore, the deletion of BglA can suppress the expression of Cbp and the deletion of Cbp can up-regulate the expression of BglA, indicating that BglA and Cbp have cross-regulation and BglA is also critical for cellobiose metabolism. These insights pave the way for both a fundamental understanding of metabolism and regulation in C. thermocellum and emphasize the importance of the degradation and utilization of polysaccharides containing β-1,3-linked glycosidic bonds in lignocellulose biorefinery.

Keywords: Biorefinery; Cellobiose; Crystal structure; Genetic deletion; Laminaribiose; β-Glucosidase.