title: Chromosomal and plasmid-encoded enzymes are required for assembly of the R3-type core oligosaccharide in the lipopolysaccharide of Escherichia coli O157:H7.
abstract: The type R3 core oligosaccharide predominates in the lipopolysaccharides from enterohemorrhagic Escherichia coli isolates including O157:H7. The R3 core biosynthesis (waa) genetic locus contains two genes, waaD and waaJ, that are predicted to encode glycosyltransferases involved in completion of the outer core. Through determination of the structures of the lipopolysaccharide core in precise mutants and biochemical analyses of enzyme activities, WaaJ was shown to be a UDP-glucose:(galactosyl) lipopolysaccharide alpha-1,2-glucosyltransferase, and WaaD was shown to be a UDP-glucose:(glucosyl)lipopolysaccharide alpha-1,2-glucosyltransferase. The residue added by WaaJ was identified as the ligation site for O polysaccharide, and this was confirmed by determination of the structure of the linkage region in serotype O157 lipopolysaccharide. The initial O157 repeat unit begins with an N-acetylgalactosamine residue in a beta-anomeric configuration, whereas the biological repeat unit for O157 contains alpha-linked N-acetylgalactosamine residues. With the characterization of WaaJ and WaaD, the activities of all of the enzymes encoded by the R3 waa locus are either known or predicted from homology data with a high level of confidence. However, when core oligosaccharide structure is considered, the origin of an additional alpha-1,3-linked N-acetylglucosamine residue in the outer core is unknown. The gene responsible for a nonstoichiometric alpha-1,7-linked N-acetylglucosamine substituent in the heptose (inner core) region was identified on the large virulence plasmids of E. coli O157 and Shigella flexneri serotype 2a. This is the first plasmid-encoded core oligosaccharide biosynthesis enzyme reported in E. coli.
authors.affiliation: Department of Microbiology, University of Guelph, Guelph, Ontario N1G 2W1, Canada.
authors.forename: Natalia A
authors.initials: NA
authors.lastname: Kaniuk
authors.forename: Evgeny
authors.initials: E
authors.lastname: Vinogradov
authors.forename: Jianjun
authors.initials: J
authors.lastname: Li
authors.forename: Mario A
authors.initials: MA
authors.lastname: Monteiro
authors.forename: Chris
authors.initials: C
authors.lastname: Whitfield
mesh_terms: D001483:Base Sequence; D002240:Carbohydrate Sequence; D002876:Chromosomes, Bacterial / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D019453:Escherichia coli O157 / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D009844:Oligosaccharides / Q000737:chemistry* / Q000378:metabolism; D009876:Operon; D010957:Plasmids / Q000235:genetics
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D008070:Lipopolysaccharides; D009844:Oligosaccharides; D016695:Glycosyltransferases