Department of Systems Biotechnology, Chung-Ang University, Anseong, 17546, Republic of Korea.
IT Convergence Materials R&BD Group, Chungcheong Regional Division, Korea Institute of Industrial Technology (KITECH), Cheonan, 31056, Republic of Korea.
Appl Microbiol Biotechnol. 2018 Aug;102(15):6515-6523. doi: 10.1007/s00253-018-9075-5. Epub 2018 May 22.
A gene encoding an endoglucanase belonging to subfamily C of glycoside hydrolase family 45 (GH45) was identified in the brown rot fungus Fomitopsis palustris and functionally expressed in Pichia pastoris. The recombinant protein displayed hydrolytic activities toward various substrates such as carboxymethyl cellulose, phosphoric acid swollen cellulose, glucomannan, lichenan, and β-glucan. In particular, the enzyme had a unique catalytic efficiency on β-1,4-glucans rather than mixed β-1,3/1,4-glucans as compared to other GH45 endoglucanases. The fungal enzyme was relatively thermostable, retaining more than 91.4% activity at 80 °C for 1 h. Site-directed mutagenesis studies revealed that the mutants N95D and D117N had significantly reduced enzymatic activities, indicating that both residues are essential for the catalytic reaction. Our study expands knowledge and understanding of the catalytic mechanism of GH45 subfamily C enzymes and also suggests that this thermostable endoglucanase from F. palustris has great potential in industrial applications.
在褐腐菌 Fomitopsis palustris 中鉴定出一个编码属于糖苷水解酶家族 45（GH45）亚家族 C 的内切葡聚糖酶的基因，并在毕赤酵母中进行了功能表达。重组蛋白对羧甲基纤维素、磷酸膨胀纤维素、葡甘露聚糖、地衣聚糖和 β-葡聚糖等各种底物表现出水解活性。与其他 GH45 内切葡聚糖酶相比，该酶对 β-1,4-葡聚糖具有独特的催化效率，而不是混合的 β-1,3/1,4-葡聚糖。真菌酶相对耐热，在 80°C 下保持超过 91.4%的活性 1 小时。定点突变研究表明，突变体 N95D 和 D117N 的酶活性显著降低，表明这两个残基对催化反应都是必需的。我们的研究扩展了对 GH45 亚家族 C 酶催化机制的认识和理解，同时表明来自 F. palustris 的这种耐热内切葡聚糖酶在工业应用中具有巨大潜力。
