A novel mechanism of β-glucosidase stimulation through a monosaccharide binding-induced conformational change

• Published in: International journal of biological macromolecules 2021-01, Vol.166, p.1188-1196
• Main Authors: Corrêa, Thamy L.R., Franco Cairo, João Paulo L., Cota, Junio, Damasio, André, Oliveira, Leandro C., Squina, Fabio M.
• Format: Article
• Language: English
• Subjects: beta-Glucosidase - chemistry; beta-Glucosidase - metabolism; Biocatalysis; Bioeconomy; Catalytic Domain; GH1; Glucose - metabolism; Kinetics; Molecular Dynamics Simulation; Monosaccharides - metabolism; Mutant Proteins - chemistry; Mutant Proteins - metabolism; Protein Binding; Protein Conformation; Thermotoga - enzymology; Thermotoga petrophila; Xylose - metabolism; β-Glucosidases
• ISSN: 0141-8130; 1879-0003
• DOI: 10.1016/j.ijbiomac.2020.11.001

It is urgent the transition from a fossil fuel-based economy to a sustainable bioeconomy based on bioconversion technologies using renewable plant biomass feedstocks to produce high chemicals, bioplastics, and biofuels. β-Glucosidases are key enzymes responsible for degrading the plant cell wall polymers, as they cleave glucan-based oligo- and polysaccharides to generate glucose. Monosaccharide-tolerant or -stimulated β-glucosidases have been reported in the past decade. Here, we describe a novel mechanism of β-glucosidase stimulation by glucose and xylose. The glycoside hydrolase 1 family β-glucosidase from Thermotoga petrophila (TpBgl1) displays a typical glucose stimulation mechanism based on an increased Vmax and decreased Km in response to glucose. Through molecular docking and dynamics analyses, we mapped putative monosaccharide binding regions (BRs) on the surface of TpBgl1. Our results indicate that after interaction with glucose or xylose at BR1 site, an adjacent loop region assumes an extended conformation, which increases the entrance to the TpBgl1 active site, improving product formation. Biochemical assays with TpBgl1 BR1 mutants, TpBgl1D49A/Y410A and TpBgl1D49K/Y410H, resulted in decreasing and abolishing monosaccharide stimulation, respectively. These mutations also impaired the BR1 looping extension responsible for monosaccharide stimulation. This study provides a molecular basis for the rational design of β-glucosidases for biotechnological applications. [Display omitted] •β-Glucosidases are key enzymes for cellulose degradation.•GH1 β-glucosidases can be monosaccharide-tolerant or -stimulated enzymes.•Docking studies found monosaccharide-binding sites (BRs) on Thermotoga petrophila TpBgl1.•Activity stimulation is related to structural change near to BR1 from TpBgl1.•This finding represents a new stimulation mechanism for GH1 β-glucosidases.