Abstract
The gene(ABK51908) from Acidothermus cellulolyticus encodes a mature protein of 484 residues with a calculated molecular mass of 53.0 kDa. Sequence analysis revealed that the protein had 59% identity to the β-glucosidases CAA82733, which belongs to glycoside hydrolase family 1(GH1). We cloned and expressed the gene in Escherichia coli BL21-Gold(DE3). The recombinant protein(AcBg) had an optimal pH and temperature of 7.0 and 70 °C, respectively. The specific activities of AcBg under optimal conditions were 290 and 33 U/mg for p-nitrophenyl-β-D-glucopyranoside(pNPG) and cellobiose, respectively. AcBg hydrolyzed the oligosaccharide sequentially from the non-reducing end to produce glucose units according to the results of HPLC analysis. AcBg showed high salt tolerance and monosaccharide-stimulation properties. Its activity rose more than 2-fold when 5 mol/L NaCl/KCl were added. The activity of the β-glucosidase was remarkably enhanced in the presence of 0.2 mol/L D-glucose(increased more than 1.9-fold), 0.1 mol/L α-methyl-D-glucose(increased more than 1.4-fold) and 1.0 mol/L D-xylose(increased more than 1.9-fold). The catalysis kinetics and structural changes in various concentrations of glucose were determined. The results indicate that glucose reduces substrate affinity and causes conformational changes.
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Supported by the National Basic Research Program of China(No.2012CB721003), the Sic-Tech Development Plan of Jilin Province, China(No.20170204040GX) and the Project of the Department of Education of Jilin Province, China(No.2016-438).
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Li, Y., Bu, M., Chen, P. et al. Characterization of a Thermophilic Monosaccharide Stimulated β-Glucosidase from Acidothermus cellulolyticus. Chem. Res. Chin. Univ. 34, 212–220 (2018). https://doi.org/10.1007/s40242-018-7408-7
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DOI: https://doi.org/10.1007/s40242-018-7408-7