期刊
CHEMISTRY-A EUROPEAN JOURNAL
卷 26, 期 3, 页码 643-649出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/chem.201903994
关键词
Bacillus subtilis lipase A; directed evolution; foldX; protein engineering; recombination
资金
- China Scholarship Council (CSC) [201604910840]
- JARA-HPC from RWTH Aachen University [JARA0169]
A main remaining challenge in protein engineering is how to recombine beneficial substitutions. Systematic recombination studies show that poorly performing variants are usually obtained after recombination of 3 to 4 beneficial substitutions. This limits researchers in exploiting nature's potential in generating better enzymes. The Computer-assisted Recombination (CompassR) strategy provides a selection guide for beneficial substitutions that can be recombined to gradually improve enzyme performance by analysis of the relative free energy of folding (Delta Delta G(fold)). The performance of CompassR was evaluated by analysis of 84 recombinants located on 13 positions of Bacillus subtilis lipase A. The finally obtained variant F17S/V54K/D64N/D91E had a 2.7-fold improved specific activity in 18.3 % (v/v) 1-butyl-3-methylimidazolium chloride ([BMIM][Cl]). In essence, the deducted CompassR rule allows recombination of beneficial substitutions in an iterative manner and empowers researchers to generate better enzymes in a time-efficient manner.
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