Enhancing the thermostability of Rhizopus oryzae lipase by combined mutation of hot-spots and engineering a disulfide bond.
Enhancing the thermostability of Rhizopus oryzae lipase by combined mutation of hot-spots and engineering a disulfide bond.
复制标题
DOI：
10.1039/c8ra07767c
复制
发表时间：
2018-12-07
期刊：
影响因子：
3.9
通讯作者：
Wu, Mian-bin
中科院分区：
文献类型：
--
作者：
Zhao, Jiong-feng;Wang, Zhe;Gao, Fei-long;Lin, Jian-ping;Yang, Li-rong;Wu, Mian-bin
Rhizopus oryzae lipase (ROL) is important because of its extreme sn-1,3-regioselectivity, but it shows poor thermostability, which severely restricts its application. In this work, the thermostability of ROL was greatly enhanced by rational design. First, several sites that may affect the thermostability of ROL were identified by multiple-sequence alignment. The half-lives of mutants V209L and D262G at 55 °C were about 4.38- and 4.2-times those of the wild-type, respectively. Then, a disulfide bond was introduced between positions 190 and 238 based on the prediction of Disulfide by Design 2, which greatly improved the thermostability of the protein. The activity of variant E190C/E238C retained about 58.2% after incubation at 55 °C for 720 min, whereas the half-life of wild type ROL was only about 11.7 min. On the basis of the results obtained by the two methods, we carried out a combined mutation. Quadruple mutant V209L/D262G/E190C/E238C was constructed and the thermostability was improved even further. The half-lives at 55 °C and 65 °C were 102.5- and 20-times those of the wild-type ROL. This improvement in thermostability will give ROL wider industrial applicability, especially in the preparation of structured lipids. The thermostability of Rhizopus oryzae lipase was greatly enhanced by combined mutation of hot-spots and engineering a disulfide bond.
登录
查看更多内容
影响因子：
6.4
作者：
Yu XW;Wang R;Zhang M;Xu Y;Xiao R
通讯作者：
Xiao R
DOI：
10.1080/07391102.2016.1206837
发表时间：
2017-01-01
影响因子：
4.4
作者：
Khan, Faez Iqbal;Nizami, Bilal;Wei, Dong-Qing
通讯作者：
Wei, Dong-Qing
影响因子：
2.6
作者：
Niu, WN;Li, ZP;Tan, TW
通讯作者：
Tan, TW
影响因子：
2.9
作者：
Hibi, Takao;Kume, Asami;Nishiya, Yoshiaki
通讯作者：
Nishiya, Yoshiaki
影响因子：
3.1
作者：
Dhara, Rupali;Dhar, Pubali;Ghosh, Mahua
通讯作者：
Ghosh, Mahua