Analysis of the role of HP0208, a phase-variable open reading frame, and its homologues HP1416 and HP0159 in the biosynthesis of Helicobacter pylori lipopolysaccharide.
The roles of the three ORFs HP0208, HP0159 and HP1416 in the biosynthesis of Helicobacter pylori 26695 LPS were investigated in this study. These ORFs represent a paralogous family of genes with homology to the Salmonella enterica serovar Typhimurium (hereafter referred to as S. typhimurium) waaJ gene, which encodes an alpha-1,2-glycosyltransferase required for core LPS biosynthesis. HP0208 contains multiple tandem repeats of the dimer 5'GA at its 5' end and its expression is predicted to be subject to phase variation. The number of 5'GA repeats present in this ORF was found to be non-permissive for the expression of HP0208 in the majority of H. pylori strains examined. To determine a role for this ORF in LPS biosynthesis a non-phase-variable, constitutively expressed variant of HP0208 was constructed and introduced into the genome of H. pylori 26695. Analysis of the LPS profile of this strain by Tricine-SDS-PAGE and immunoblotting with anti-Lewis Y antigen (Le(y)) mAbs confirmed a role for HP0208 in the biosynthesis of core LPS. A role for HP0159 and HP1416 in the biosynthesis of core LPS was also established. Although homologous to waaJ, H. pylori HP0208, HP0159 and HP1416 failed to complement an S. typhimurium waaJ mutant, suggesting that these ORFs encode functionally different enzymes.
Langdon
Rebecca
R
University of Manchester, Faculty of Life Sciences, 1.800 Stopford Building, Oxford Road, Manchester M13 9PT, UK.
Craig
Jane E
JE
University of Manchester, Faculty of Life Sciences, 1.800 Stopford Building, Oxford Road, Manchester M13 9PT, UK.
Goldrick
Marie
M
University of Manchester, Faculty of Life Sciences, 1.800 Stopford Building, Oxford Road, Manchester M13 9PT, UK.
Houldsworth
Rebecca
R
University of Manchester, Faculty of Life Sciences, 1.800 Stopford Building, Oxford Road, Manchester M13 9PT, UK.
High
Nicola J
NJ
University of Manchester, Faculty of Life Sciences, 1.800 Stopford Building, Oxford Road, Manchester M13 9PT, UK.
D001426:Bacterial Proteins / Q000502:physiology; D003001:Cloning, Molecular; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000502:physiology*; D016480:Helicobacter pylori / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000096:biosynthesis* / Q000737:chemistry; D016366:Open Reading Frames / Q000502:physiology*
D001426:Bacterial Proteins; D008070:Lipopolysaccharides