title: The crystal structure of 1-D-myo-inosityl 2-acetamido-2-deoxy-alpha-D-glucopyranoside deacetylase (MshB) from Mycobacterium tuberculosis reveals a zinc hydrolase with a lactate dehydrogenase fold.
abstract: Mycothiol (1-D-myo-inosityl 2-(N-acetyl-L-cysteinyl)amido-2-deoxy-alpha-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 Zn2+ 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 beta-sheets and two alpha-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 Zn2+ 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 Zn2+ 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.
authors.affiliation: Canadian Institutes of Health Research, Group in Protein Structure and Function, Department of Biochemistry, Faculty of Medicine, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
authors.forename: Jason T
authors.initials: JT
authors.lastname: Maynes
authors.forename: Craig
authors.initials: C
authors.lastname: Garen
authors.forename: Maia M
authors.initials: MM
authors.lastname: Cherney
authors.forename: Gerald
authors.initials: G
authors.lastname: Newton
authors.forename: Dorit
authors.initials: D
authors.lastname: Arad
authors.forename: Yossef
authors.initials: Y
authors.lastname: Av-Gay
authors.forename: Robert C
authors.initials: RC
authors.lastname: Fahey
authors.forename: Michael N G
authors.initials: MN
authors.lastname: James
mesh_terms: D000581:Amidohydrolases / Q000737:chemistry*; D001426:Bacterial Proteins; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006867:Hydrolases / Q000737:chemistry; D007770:L-Lactate Dehydrogenase / Q000737:chemistry; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014867:Water / Q000737:chemistry; D015032:Zinc / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D014867:Water; D007770:L-Lactate Dehydrogenase; D006867:Hydrolases; D000581:Amidohydrolases; C417275:N-acetyl-1-D-inosityl-2-amino-2-deoxy-alpha-D-glucopyranoside deacetylase; D015032:Zinc