Abstract
A β-glucosidase gene (bsbgl1a) from Bacillus sp. CGMCC 1.16541 was expressed in Escherichia coli BL21 and subsequently characterized. The amino acid sequence shared 83.64% identity with β-glucosidase (WP_066390903.1) from Fictibacillus phosphorivorans. The recombinant β-glucosidase (BsBgl1A) had a molecular weight of 52.2 kDa and could hydrolyze cellobiose, cellotriose, cellotetrose, p-nitrophenyl-β-d-glucopyranoside (pNPG), and p-nitrophenyl-β-d-xylopyranoside (pNPX). Optimal activity for BsBgl1A was recorded at 45 °C with a pH between 5.6 and 7.6, and 100% of its activity was maintained after a 24 h incubation between pH 4 and 9. Kinetic characterization revealed an enzymatic turnover (Kcat) of 616 ± 2 s−1 (with cellobiose) and 3.5 ± 0.1 s−1 (with p-nitrophenyl-β-d-glucopyranoside). Interestingly, the recombinant enzyme showed cupric ion (Cu2+), sodium dodecyl sulfate (SDS) and alcohol tolerance at 10 mM for Cu2+ and 10% for both SDS and alcohol. Additionally, BsBgl1A had high tolerance for glucose (Ki = 2095 mM), which is an extremely desirable feature for industrial applications. Following the addition of BsBgl1A (0.05 mg/ml) to a commercial cellulase reaction system, glucose yields from sugarcane bagasse increased 100% after 1 day at 45 °C. This work identifies a Cu2+, SDS, alcohol, and glucose tolerant GH1 β-glucosidase with potential applications in the hydrolysis of cellulose for the bioenergy industry.
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Acknowledgements
This research was supported by the National Natural Sciences Foundation of China Regional Program (Grant Nos. 31660015 and 31860243), Yunnan Applied Basic Research Projects (Grant Nos. 2017FB024 and 2017FH001-032) and the Chinese Postdoctoral Science Foundation (Grant No. 2017M622861).
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Y.R.Y., L.Q.Y. and W.J.L. conceived the study. P.S. and F.L.Y. were responsible for all PCR and gene cloning. R.F.Y. cultured strains and collected samples. H.Y.L. conducted protein separation. Z.L.L. measured enzymatic activity. T.L. performed data analysis and phylogenetic mapping. Y.R.Y., P.S., L.Q.Y., and W.J.L. wrote the manuscript. All authors discussed the results and commented on the manuscript. All authors read and approved the final manuscript.
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Yin, YR., Sang, P., Yang, FL. et al. Characterization of a Cu2+, SDS, alcohol and glucose tolerant GH1 β-glucosidase from Bacillus sp. CGMCC 1.16541. Antonie van Leeuwenhoek 113, 1467–1477 (2020). https://doi.org/10.1007/s10482-020-01455-w
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DOI: https://doi.org/10.1007/s10482-020-01455-w