Discovery and Structural Analysis to Improve the Enantioselectivity of Hydroxynitrile Lyase from <i>Parafontaria laminata</i> Millipedes for (<i>R</i>)-2-Chloromandelonitrile Synthesis
Aem Nuylert, Makoto Nakabayashi, Takuya Yamaguchi, Yasuhisa Asano
Abstract
, although the forming binding cavity was different. In order to improve the catalytic activity and stereoselectivity, a computational structure-guided directed evolution approach was performed by an enzyme-substrate docking simulation at all of the residues that were exposed on the surface of the active site. The PlamHNL-N85Y mutant showed higher conversion (91% conversion with 98.2% ee of the product) than the wild type (76% conversion with 90% ee of the product) at pH 3.5 and 25 °C for 30 min of incubation. This study shows the diversity of millipede HNLs and reveals the molecular basis for improvement of the activity and stereoselectivity of the wild-type HNL to increase the reaction rate and enantioselectivity in the synthesis of 2-chloromandelonitrile.