Abstract:A β-glucosidase gene SmBgl3A, which might play a key role in cellobiose degradation and utilization, was screened from Sporocytophaga sp. CX11 through the analysis and comparison of expression level. The gene contains 2 283 bp and encodes 760 amino acids. The encoded protein of SmBgl3A has 46.76% homology with the β-glucosidase BoGH3B from Bacteroides ovatus. The gene SmBgl3A was heterologously expressed in Escherichia coli. The electrophoretically pure recombinant protein SmBgl3A was achieved by the purification of crude enzymes using Ni-NTA affinity chromatography, and the relative molecular weight of recombinant protein was consistent with the theoretical value (81.2×104). Characterization of enzymatic properties showed that the optimum temperature and pH of SmBgl3A were 35 ℃ and 6.5, respectively. When the pH value was 5.0~7.0 and the temperature was below 40 ℃, SmBgl3A was stable. In addition, SmBgl3A showed good NaCl tolerance. The optimal substrate for SmBgl3A was pNPG, and the specific enzyme activity with pNPG as substrate was 14.74 U/mg. Km and kcat of SmBgl3A towards pNPG were 10.88 mmol/L and 46.72 s-1, respectively. SmBgl3A is a novel enzyme with excellent enzymatic properties, further expanding the kinds of β-glucosidase and laying a foundation for the enzyme application in cellulose degradation, food industry and other fields.
