Directed Evolution and Resolution Mechanism of 1, 3-Propanediol Oxidoreductase from Klebsiella pneumoniae toward Higher Activity by Error-Prone PCR and Bioinformatics
Year: 2015
Type: article
Abstract: 1, 3-propanediol oxidoreductase (PDOR) is a key enzyme in glycerol bioconversion to 1,3-propanediol (1, 3-PD) which is a valuable chemical and one of the six new petrochemical products. We used error-prone PCR and activity screening to identify mutants of Klebsiella pneumoniae (K. pneumoniae) PDOR with improved activity. The activity of one of the identified mutants, PDOR'-24, which includes a single mutation, A199S, was 48 U/mg, 4.9 times that of the wild-type enzyme. Molecular docking was performed to analyze the identified mutants; and amino acids S103, H271, N366, D106, N262 and D364 were predicted to bond with NADH. The origins of the improved activity of PDOR'-24, as well as three other mutants were analyzed by simulating the interaction mechanism of the mutants with the substrate and coenzyme, respectively. This research provides useful information about the use of safranine O plate screening for the directed evolution of oxidoreductases, identifies interesting sites for improving PDOR activity, and demonstrates the utility of using molecular docking to analyze the interaction mechanism of the mutants with the substrate and coenzyme, respectively.
Source: PLoS ONE
Authors: Wei Jiang, Yuan Zhuang, Shizhen Wang, Baishan Fang
Institutions: Xiamen University, Huaqiao University
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,382
OpenAlex ID: W1934869815