Structure-based engineering of ω-transaminase for enhanced catalytic efficiency toward (R)-(+)-1-(1-naphthyl)ethylamine synthesis

Abstract

ω-Transaminase is a promising enzyme for the synthesis of chiral (R)-(+)-1-(1-naphthyl)ethylamine, a key pharmaceutical intermediate for many chiral drugs. However, the low catalytic efficiency of ω-transaminase toward 1-acetonaphthone substrate severely limits the industrial synthesis of (R)-(+)-1-(1-naphthyl)ethylamine. Herein, a structure-based engineering strategy combining in silico and in vitro studies was developed to engineer a ω-transaminase from Arthrobacter sp. for improved catalytic efficiency toward 1-acetonaphthone. On basis of homologous modeling and molecular docking analysis, alanine scanning, saturation and combinatorial mutagenesis of G136, V199, and S223 residues were performed. Notably, double-variant V199W/S223P showed 8.5-fold increased catalytic efficiency. In addition, triple-variant G136I/V199W/S223 P showed improved thermostability with 9.1 % and 6.2 % enhanced half-life at 30 °C and 40 °C, respectively. Possible mechanisms were elucidated through molecular docking and molecular dynamics simulation. Enhanced catalytic efficiency could be attributed to increased interactions between variants and 1-acetonaphthone. Reduced flexibility of loop 130–139 may be responsible for the improved thermostability of G136I/V199W/S223P. Our results provides useful information for engineering ω-transaminase toward (R)-(+)-1-(1-naphthyl)ethylamine production.