Role of Ser216 in the mechanism of action of membrane-bound lytic transglycosylase B: further evidence for substrate-assisted catalysis.
Lytic transglycosylases cleave the beta-(1-->4)-glycosidic bond in the bacterial cell wall heteropolymer peptidoglycan between the N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc) residues with the concomitant formation of a 1,6-anhydromuramoyl residue. Based on sequence alignments, Ser216 in Pseudomonas aeruginosa membrane-bound lytic transglycosylase B (MltB) was targeted for replacement with alanine to delineate its role in the enzyme's mechanism of action. The specific activity of the Ser216-->Ala MltB derivative was less than 12% of that for the wild-type enzyme, while its substrate binding affinity remained virtually unaltered. These data are in agreement with a role of Ser216 in orienting the N-acetyl group on MurNAc at the -1 subsite of MltB for its participation in a substrate-assisted mechanism of action.
Reid
Christopher W
CW
Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario, Canada N1G 2W1.
Legaree
Blaine A
BA
Clarke
Anthony J
AJ
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001665:Binding Sites; D002384:Catalysis; D016695:Glycosyltransferases / Q000737:chemistry* / Q000235:genetics; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D010457:Peptidoglycan / Q000737:chemistry / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000201:enzymology; D012694:Serine / Q000737:chemistry*
D001426:Bacterial Proteins; D008565:Membrane Proteins; D010457:Peptidoglycan; D012694:Serine; D016695:Glycosyltransferases