College of Biological Sciences and Biotechnology, Beijing Forestry University, Beijing 100083, People's Republic of China; Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, People's Republic of China.
Key Laboratory for Feed Biotechnology of the Ministry of Agriculture, Feed Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, People's Republic of China.
Bioresour Technol. 2016 Oct;218:279-85. doi: 10.1016/j.biortech.2016.06.094. Epub 2016 Jun 25.
The aim of this work was to study the contribution of the N-terminal structure to cellulase catalytic performance. A wild-type cellulase (BaCel5) of glycosyl hydrolase (GH) family 5 from Bispora antennata and two hybrid enzymes (BaCel5(127) and BaCel5(167)) with replacement of the N-terminal (βα)3 (127 residues) or (βα)4 (167 residues)-barrel with the corresponding sequences of TeEgl5A from Talaromyces emersonii were produced in Pichia pastoris and biochemically characterized. BaCel5 exhibited optimal activity at pH 5.0 and 50°C but had low catalytic efficiency (25.4±0.8mLs(-1)mg(-1)). In contrast, BaCel5(127) and BaCel5(167) showed similar enzymatic properties but improved catalytic performance. When using CMC-Na, barley β-glucan, lichenan, and cellooligosaccharides as substrates, BaCel5(127) and BaCel5(167) had increased specific activities and catalytic efficiencies by ∼1.8-6.7-fold and ∼1.0-4.7-fold, respectively. The catalytic efficiency of BaCel5(167) was even higher than that of parental proteins. The underlying mechanism was analyzed by molecular docking and molecular dynamic simulation.
本工作旨在研究 N 端结构对纤维素酶催化性能的贡献。从双孢蚁巢菌中分离到糖苷水解酶（GH）家族 5 的野生型纤维素酶（BaCel5）和两种杂合酶（BaCel5（127）和 BaCel5（167）），它们的 N 端（βα）3（127 个残基）或（βα）4（167 个残基）-桶分别被来自塔宾曲霉的 TeEgl5A 的相应序列取代，在巴斯德毕赤酵母中表达并进行了生化特性分析。BaCel5 在 pH5.0 和 50°C 时表现出最佳活性，但催化效率较低（25.4±0.8mLs(-1)mg(-1)）。相比之下，BaCel5（127）和 BaCel5（167）表现出相似的酶学性质，但催化性能得到了提高。当以 CMC-Na、大麦β-葡聚糖、地衣聚糖和纤维寡糖为底物时，BaCel5（127）和 BaCel5（167）的比活性和催化效率分别提高了约 1.8-6.7 倍和约 1.0-4.7 倍。BaCel5（167）的催化效率甚至高于亲本蛋白。通过分子对接和分子动力学模拟分析了其潜在机制。
Appl Environ Microbiol. 2019-2-20
Appl Environ Microbiol. 2017-3-2
Biotechnol Biofuels. 2018-3-22
Appl Environ Microbiol. 2019-2-20
Zotero 插件