RRC ID 54564
Author Honda Y, Nakano S, Ito S, Dadashipour M, Zhang Z, Kawarabayasi Y.
Title Improvement of ST0452 N-Acetylglucosamine-1-Phosphate Uridyltransferase Activity by the Cooperative Effect of Two Single Mutations Identified through Structure-Based Protein Engineering.
Journal Appl. Environ. Microbiol.
Abstract We demonstrated that the enzymatic activity of a thermostable protein was over 4 times higher than that of the wild-type protein following substitution of a single amino acid, without affecting its thermostability. The three-dimensional structure of the improved mutant protein complexed with substrate was determined. The same overall structure and interaction between the substituted residue and the GlcNAc substrate as observed in the well-characterized bacterial enzyme suggested that the substitution of Tyr at position 97 by Asn might slightly change the interaction. This subtle change in the interaction might potentially increase the GlcNAc-1-P UTase activity of the mutant protein. These observations indicated that a drastic change in the structure of a natural thermostable enzyme is not necessary to increase its activity; a subtle change in the interaction with the substrate might be sufficient. Cooperative effects were observed in the appropriate double mutant protein. This work provides useful information for the future engineering of natural enzymes.
Volume 84(24)
Published 2018-12-15
DOI 10.1128/AEM.02213-18
PII AEM.02213-18
PMID 30291121
PMC PMC6275352
IF 3.633
Resource
DNA material Thermophile Gene Expression Plasmid (RDB15969)