The Crystal Structure of 1-D-myo-Inosityl 2-Acetamido-2-deoxy-α-D-glucopyranoside Deacetylase (MshB) from Mycobacterium tuberculosis Reveals a Zinc Hydrolase with a Lactate Dehydrogenase Fold

• Published in: The Journal of biological chemistry 2003-11, Vol.278 (47), p.47166
• Main Authors: Jason T. Maynes, Craig Garen, Maia M. Cherney, Gerald Newton, Dorit Arad, Yossef Av-Gay, Robert C. Fahey, Michael N. G. James
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M308914200

Mycothiol (1- d - myo -inosityl 2-( N -acetyl- l -cysteinyl)amido-2-deoxy-α- d -glucopyranoside, MSH or AcCys-GlcN-inositol (Ins)) is the major reducing agent in actinomycetes, including Mycobacterium tuberculosis . The biosynthesis of MSH involves a deacetylase that removes the acetyl group from the precursor GlcNAc-Ins to yield GlcN-Ins. The deacetylase (MshB) corresponds to Rv1170 of M. tuberculosis with a molecular mass of 33,400 Da. MshB is a Zn 2+ metalloprotein, and the deacetylase activity is completely dependent on the presence of a divalent metal cation. We have determined the x-ray crystallographic structure of MshB, which reveals a protein that folds in a manner resembling lactate dehydrogenase in the N-terminal domain and a C-terminal domain consisting of two β-sheets and two α-helices. The zinc binding site is in the N-terminal domain occupying a position equivalent to that of the NAD + co-factor of lactate dehydrogenase. The Zn 2+ is 5 coordinate with 3 residues from MshB (His-13, Asp-16, His-147) and two water molecules. One water would be displaced upon binding of substrate (GlcNAc-Ins); the other is proposed as the nucleophilic water assisted by the general base carboxylate of Asp-15. In addition to the Zn 2+ providing electrophilic assistance in the hydrolysis, His-144 imidazole could form a hydrogen bond to the oxyanion of the tetrahedral intermediate. The extensive sequence identity of MshB, the deacetylase, with mycothiol S -conjugate amidase, an amide hydrolase that mediates detoxification of mycothiol S -conjugate xenobiotics, has allowed us to construct a faithful model of the catalytic domain of mycothiol S -conjugate amidase based on the structure of MshB.