Campylobacter jejuni 81-176 lipooligosaccharide (LOS) is composed of two covalently linked domains: lipid A, a hydrophobic anchor, and a nonrepeating core oligosaccharide, consisting of an inner and outer core region. We report the isolation and characterization of the deepest rough C. jejuni 81-176 mutant by insertional mutagenesis into the waaC gene, encoding heptosyltransferase I that catalyzes the transfer of the first l-glycero-d-manno-heptose residue to 3-deoxy-d-manno-octulosonic residue (Kdo)-lipid A. Tricine gel electrophoresis, followed by silver staining, showed that site-specific mutation in the waaC gene resulted in the expression of a severely truncated LOS compared to wild-type strain 81-176. Gas-liquid chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy showed that the waaC LOS species lacked all sugars distal to Kdo-lipid A. Parallel structural studies of the capsular polysaccharides of the wild-type strain 81-176 and waaC mutant revealed loss of the 3-O-methyl group in the waaC mutant. Complementation of the C. jejuni mutant by insertion of the wild-type C. jejuni waaC gene into a chromosomal locus resulted in LOS and capsular structures identical to those expressed in the parent strain. We also report here the presence of O-methyl phosphoramidate in wild-type strain 81-176 capsular polysaccharide.
capsular polysaccharide, Campylobacter jejuni, core region, heptosyltransferase I
NCBI PubMed ID: 16621820Publication DOI: 10.1128/JB.188.9.3273-3279.2006Journal NLM ID: 2985120RPublisher: American Society for Microbiology
Correspondence: guerryp@nmrc.navy.mil
Institutions: Department of Chemistry, University of Guelph, Guelph, ON N1G 2W1, Canada, Enteric Diseases Department, Naval Medical Research Center, 503 Robert Grant Avenue, Silver Spring, MD 20910, and Enteric Diseases Department, Naval Medical Research Center, Silver Spring, Maryland 20910
Methods: GLC-MS, FAB-MS, NMR, SDS-PAGE, GLC, composition analysis, genetic methods
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