Structural insights into a bacterial β-glucosidase capable of degrading sesaminol triglucoside to produce sesaminol: toward the understanding of the aglycone recognition mechanism by the C-terminal lid domain

Abstract

The sesaminol triglucoside (STG)-hydrolyzing β-glucosidase from Paenibacillus sp. (PSTG1), which belongs to glycoside hydrolase family 3 (GH3), is a promising catalyst for the industrial production of sesaminol. We determined the X-ray crystal structure of PSTG1 with bound glycerol molecule in the putative active site. PSTG1 monomer contained typical three domains of GH3 with the active site in domain 1 (TIM barrel). In addition, PSTG1 contained an additional domain (domain 4) at the C-terminus that interacts with the active site of the other protomer as a lid in the dimer unit. Interestingly, the interface of domain 4 and the active site forms a hydrophobic cavity probably for recognizing the hydrophobic aglycone moiety of substrate. The short flexible loop region of TIM barrel was found to be approaching the interface of domain 4 and the active site. We found that n-heptyl-β-D-thioglucopyranoside detergent acts as an inhibitor for PSTG1. Thus, we propose that the recognition of hydrophobic aglycone moiety is important for PSTG1-catalyzed reactions. Domain 4 might be a potential target for elucidating the aglycone recognition mechanism of PSTG1 as well as for engineering PSTG1 to create a further excellent enzyme to degrade STG more efficiently to produce sesaminol.

Keywords: glycoside hydrolase family 3; sesaminol triglucoside; β-glucosidase.Abbreviations: STG, sesaminol triglucoside; PSTG1, STG-hydrolyzing β-glucosidase from Paenibacillus sp; GH3, Glycoside Hydrolase Family 3; TIM, Triosephosphate isomerase, Fn-III, Fibronectin type III; 2-SDG, 2-O-(β-D-glucopyranosyl)-β-D-glucopyranosylsesaminol; 6-SDG, 6-O-(β-D-glucopyranosyl)-β-D-glucopyranosylsesaminol; SMG, β-D-glucopyranosylsesaminol; HTG, n-Heptyl-beta-D-thioglucopyranoside; OTG, n-Octyl-β-D-glucoside; pNP-β-Glc, p-Nitrophenyl-β-D-glucopyranoside.