Structure of the TDP-epi-vancosaminyltransferase GtfA from the chloroeremomycin biosynthetic pathway.
During the biosynthesis of the vancomycin-class antibiotic chloroeremomycin, TDP-epi-vancosaminyltransferase GtfA catalyzes the attachment of 4-epi-vancosamine from a TDP donor to the beta-OHTyr-6 of the aglycone cosubstrate. Glycosyltransferases from this pathway are potential tools for the combinatorial design of new antibiotics that are effective against vancomycin-resistant bacterial strains. These enzymes are members of the GT-B glycosyltransferase superfamily, which share a homologous bidomain topology. We present the 2.8-A crystal structures of GtfA complexes with vancomycin and the natural monoglycosylated peptide substrate, representing the first direct observation of acceptor substrate binding among closely related glycosyltransferases. The acceptor substrates bind to the N-terminal domain such that the aglycone substrate's reactive hydroxyl group hydrogen bonds to the side chains of Ser-10 and Asp-13, thus identifying these as residues of potential catalytic importance. As well as an open form of the enzyme, the crystal structures have revealed a closed form in which a TDP ligand is bound at a donor substrate site in the interdomain cleft, thereby illustrating not only binding interactions, but the conformational changes in the enzyme that accompany substrate binding.
Mulichak
Anne M
AM
Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI 48824, USA.
Losey
Heather C
HC
Lu
Wei
W
Wawrzak
Zdzislaw
Z
Walsh
Christopher T
CT
Garavito
R Michael
RM
D001224:Aspartic Acid / Q000737:chemistry; D001426:Bacterial Proteins / Q000378:metabolism; D001665:Binding Sites; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D016695:Glycosyltransferases / Q000378:metabolism; D006859:Hydrogen / Q000737:chemistry; D008956:Models, Chemical; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012694:Serine / Q000737:chemistry; D013379:Substrate Specificity; D000637:Transaminases / Q000737:chemistry* / Q000378:metabolism; D014640:Vancomycin / Q000031:analogs & derivatives* / Q000096:biosynthesis*
D001426:Bacterial Proteins; C112142:chloroeremomycin; D001224:Aspartic Acid; D012694:Serine; D014640:Vancomycin; D006859:Hydrogen; D016695:Glycosyltransferases; C477600:TDP-epi-vancosaminyltransferase; D000637:Transaminases