Active-Site Engineering of ω-Transaminase for Production of Unnatural Amino Acids Carrying a Side Chain Bulkier than an Ethyl Substituent.

ABSTRACT: ω-Transaminase (ω-TA) is a promising enzyme for use in the production of unnatural amino acids from keto acids using cheap amino donors such as isopropylamine. The small substrate-binding pocket of most ω-TAs permits entry of substituents no larger than an ethyl group, which presents a significant challenge to the preparation of structurally diverse unnatural amino acids. Here we report on the engineering of an (S)-selective ω-TA from Ochrobactrum anthropi (OATA) to reduce the steric constraint and thereby allow the small pocket to readily accept bulky substituents. On the basis of a docking model in which L-alanine was used as a ligand, nine active-site residues were selected for alanine scanning mutagenesis. Among the resulting variants, an L57A variant showed dramatic activity improveme

SUBMITTER: Han SW 

PROVIDER: S-EPMC4579442 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature