Department of Life Science and Biotechnology, Nanyang Normal University, Nanyang, 473000, P.R. China.
State Key Laboratory of Cotton Biology, Henan Key Laboratory of Plant Stress Biology, School of Life Sciences, Henan University, Kaifeng, 475004, China.
Sci Rep. 2017 Nov 1;7(1):14850. doi: 10.1038/s41598-017-14073-w.
β-glucosidases catalyze the final step of cellulose hydrolysis and are essential in cellulose degradation. A β-glucosidase gene, cen502, was identified and isolated from a metagenomic library from Bursaphelenchus xylophilus via functional screening. Analyses indicated that cen502 encodes a 465 amino acid polypeptide that contains a catalytic domain belonging to the glycoside hydrolase family 1 (GH1). Cen502 was heterologously expressed, purified, and biochemically characterized. Recombinant Cen502 displayed optimum enzymatic activity at pH 8.0 and 38 °C. The enzyme had highest specific activity to p-nitrophenyl-β-D-glucopyranoside (pNPG; 180.3 U/mg) and had K and V values of 2.334 mol/ml and 9.017 μmol/min/mg, respectively. The addition of Fe and Mn significantly increased Cen502 β-glucosidase activity by 60% and 50%, respectively, while 10% and 25% loss of β-glucosidase activity was induced by addition of Pb and K, respectively. Cen502 exhibited activity against a broad array of substrates, including cellobiose, lactose, salicin, lichenan, laminarin, and sophorose. However, Cen502 displayed a preference for the hydrolysis of β-1,4 glycosidic bonds rather than β-1,3, β-1,6, or β-1,2 bonds. Our results indicate that Cen502 is a novel β-glucosidase derived from bacteria associated with B. xylophilus and may represent a promising target to enhance the efficiency of cellulose bio-degradation in industrial applications.
β-葡萄糖苷酶催化纤维素水解的最后一步，是纤维素降解的关键酶。本研究通过功能筛选，从松材线虫共生菌宏基因组文库中鉴定并分离到一个β-葡萄糖苷酶基因 cen502。序列分析表明，cen502 编码一个 465 个氨基酸的多肽，包含糖苷水解酶家族 1（GH1）的催化结构域。cen502 被异源表达、纯化，并进行了生化特性分析。重组 Cen502 在 pH8.0 和 38°C 时表现出最佳酶活性。该酶对 p-硝基苯-β-D-葡萄糖苷（pNPG）的比活最高（180.3 U/mg），K 和 V 分别为 2.334mol/ml 和 9.017 μmol/min/mg。添加 Fe 和 Mn 可分别使 Cen502 的β-葡萄糖苷酶活性显著提高 60%和 50%，而添加 Pb 和 K 则分别导致酶活损失 10%和 25%。cen502 对多种底物表现出活性，包括纤维二糖、乳糖、水杨苷、昆布多糖、麦芽寡糖和槐糖。然而，cen502 对β-1,4 糖苷键的水解表现出偏好，而对β-1,3、β-1,6 或β-1,2 键的水解则较弱。本研究结果表明， Cen502 是一种新型的β-葡萄糖苷酶，来源于与松材线虫共生的细菌，可能是提高纤维素生物降解效率在工业应用中的有前途的靶点。
