College of Agriculture and Biological Science, Dali University, Dali, 671003, People's Republic of China.
Xiaguan No. 1 Middle School, Dali, 671000, People's Republic of China.
Antonie Van Leeuwenhoek. 2020 Oct;113(10):1467-1477. doi: 10.1007/s10482-020-01455-w. Epub 2020 Aug 3.
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 (with cellobiose) and 3.5 ± 0.1 s (with p-nitrophenyl-β-D-glucopyranoside). Interestingly, the recombinant enzyme showed cupric ion (Cu), sodium dodecyl sulfate (SDS) and alcohol tolerance at 10 mM for Cu 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 Cu, SDS, alcohol, and glucose tolerant GH1 β-glucosidase with potential applications in the hydrolysis of cellulose for the bioenergy industry.
从芽孢杆菌 CGMCC 1.16541 中表达了一个β-葡萄糖苷酶基因（bsbgl1a），并在大肠杆菌 BL21 中进行了后续的特性分析。该氨基酸序列与来自假黄单胞菌（Fictibacillus phosphorivorans）的β-葡萄糖苷酶（WP_066390903.1）具有 83.64%的同源性。重组β-葡萄糖苷酶（BsBgl1A）的分子量为 52.2 kDa，能够水解纤维二糖、纤维三糖、纤维四糖、对硝基苯基-β-D-吡喃葡萄糖苷（pNPG）和对硝基苯基-β-D-木吡喃糖苷（pNPX）。BsBgl1A 的最适活性在 45°C 下 pH 值在 5.6 到 7.6 之间，在 pH 值 4 到 9 之间孵育 24 小时后，其活性仍保持 100%。动力学特征表明其酶转化（Kcat）值为 616±2 s（用纤维二糖）和 3.5±0.1 s（用对硝基苯基-β-D-吡喃葡萄糖苷）。有趣的是，该重组酶在 10 mM 的铜离子（Cu）、10%的十二烷基硫酸钠（SDS）和酒精中具有耐受性，在 10 mM 的 Cu 和 10%的 SDS 和酒精中仍保持 100%的活性。此外，BsBgl1A 对葡萄糖（Ki=2095 mM）具有高耐受性，这是工业应用中非常理想的特性。在向商业纤维素酶反应体系中添加 0.05 mg/ml 的 BsBgl1A 后，在 45°C 下 1 天后，甘蔗渣的葡萄糖产量增加了 100%。这项工作鉴定了一种 Cu、SDS、酒精和葡萄糖耐受的 GH1 β-葡萄糖苷酶，在纤维素水解用于生物能源工业中具有潜在的应用价值。
