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Issue 17, 2017
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Biochemical studies of inositol N-acetylglucosaminyltransferase involved in mycothiol biosynthesis in Corynebacterium diphtheria

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Abstract

Mycothiol (MSH) is the predominant low molecular weight thiol produced by actinomycetes, and it plays a pivotal role in the bacterial detoxication process. 1L-myo-Inositol-1-phosphate (1L-Ins-1-P) α-N-acetylglucosaminyltransferase (GlcNAc-T), known as MshA, is the only glycosyltransferase involved in MSH biosynthesis. In this work, the MshA from Corynebacterium diphtheria, named as CdMshA, was expressed, purified, and studied in detail. Its enzymatic activity to transfer GlcNAc to 1L-Ins-1-P was confirmed by the isolation and rigorous characterization of its reaction product 3-phospho-1-D-myo-inositol-2-acetamido-2-deoxy-α-D-glucopyranoside. CdMshA was shown to accept only UDP-GlcNAc and 1L-Ins-1-P as its substrates among various tested glycosyl donors, such as UDP-GlcNAc, UDP-Gal, UDP-Glc, UDP-GalNAc and UDP-GlcA, and glycosyl acceptors, such as myo-inositol, 1L-Ins-1-P and 1D-Ins-1-P. The results have demonstrated the strict substrate selectivity of CdMshA. Furthermore, its reaction kinetics with UDP-GlcNAc and 1L-Ins-1-P as substrates were characterized, while site-directed mutagenesis of CdMshA disclosed that its amino acid residues N28, K81 and R157 were essential for its enzymatic activity.

Graphical abstract: Biochemical studies of inositol N-acetylglucosaminyltransferase involved in mycothiol biosynthesis in Corynebacterium diphtheria

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Supplementary files

Article information


Submitted
04 Mar 2017
Accepted
05 Apr 2017
First published
05 Apr 2017

Org. Biomol. Chem., 2017,15, 3775-3782
Article type
Paper

Biochemical studies of inositol N-acetylglucosaminyltransferase involved in mycothiol biosynthesis in Corynebacterium diphtheria

Y. Guo, L. Wang, J. Guo, G. Gu and Z. Guo, Org. Biomol. Chem., 2017, 15, 3775
DOI: 10.1039/C7OB00555E

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