High-Throughput Screening and Directed Evolution of Methionine Adenosyltransferase from Escherichia Coli

Overview

Journal Appl Biochem Biotechnol

Specialties Biochemistry
Biotechnology

Date 2023 Jan 18

PMID 36652094

Authors

Chenqi Cao

Kaili Nie

Haijun Xu

Luo Liu

Affiliations

Soon will be listed here.

Abstract

S-adenosyl-L-methionine (SAM) is the active form of methionine, which participates in various metabolic reactions and plays a vital role. It is mainly used as a precursor by three key metabolic pathways: trans-methylation, trans-sulfuration, and trans-aminopropylation. Methionine adenosyltransferase (MAT) is the only enzyme to produce SAM from methionine and ATP. However, there is no efficient and accurate method for high-throughput detection of SAM, which is the major obstacles of directed evolution campaigns for MAT. Herein, we established a colorimetric method for directed evolution of MAT based on detecting SAM by using glycine oxidase and glycine/sarcosine N-methyltransferase enzyme. Screening of MAT libraries revealed variant I303V/Q22R with 2.13-fold improved activity towards SAM in comparison to the wild type. Molecular dynamic simulation indicates that the loops more flexible and more conducive to SAM release.

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