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The putative neuraminyllactose-binding hemagglutinin HpaA of Helicobacter pylori CCUG 17874 is a lipoprotein.

ABSTRACT: The ability of certain strains of Helicobacter pylori to cause sialic acid-sensitive agglutination of erythrocytes has been attributed to the HpaA protein (D.G. Evans, T.K. Karjalainen, D. J. Evans, Jr., D. Y. Graham, and C.H. Lee, J. Bacteriol. 175:674-683, 1993), the gene for which has been cloned and sequenced. On the basis of the hydropathy plot of HpaA and the presence of a potential lipoprotein signal sequence and modification site, and because of the similarities of these features with those of the cell envelope lipoprotein Lpp20 of H. pylori, we examined the possibility that HpaA was also a lipoprotein. Posttranslational processing of the HpaA protein expressed by the cloned gene was sensitive to globomycin, an inhibitor of the lipoprotein-specific signal peptidase II. Antibodies raised to the putative sialic acid-binding region of HpaA failed to bind to the surface of H. pylori cells in immunoelectron microscopy but instead were observed to have labeled the cytoplasm when thin sections were examined. This antibody recognized a 29,000-M(r) protein in Western blots (immunoblots) of cell extracts of H. pylori and Escherichia coli cells expressing the cloned hpaA gene. Determination of the sequence of hpaA from strain CCUG 17874 indicated significant differences from that determined by Evans and coworkers in the above-mentioned study, including extension of the gene into the open reading frame 3 downstream of hpaA to produce a protein with an M(r) of 26,414. Localization of HpaA indicated that it was predominantly located in the cytoplasmic fraction of the cell in both E. coli and H. pylori. HpaA was not observed in the sarkosyl-insoluble outer membrane fraction. An isogenic mutant generated by insertional inactivation of hpaA was unaffected in its ability to bind four different human cell lines as well as fixed sections of gastric tissue and had hemagglutination properties identical to those of the wild type. The data collectively suggest that HpaA is a nonessential lipoprotein internal to the H. pylori cell and that it is not involved in adhesion.

SUBMITTER: O'Toole PW

PROVIDER: S-EPMC177441 | biostudies-other | 1995 Nov

REPOSITORIES: biostudies-other

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The putative neuraminyllactose-binding hemagglutinin HpaA of Helicobacter pylori CCUG 17874 is a lipoprotein.

O'Toole P W PW Janzon L L Doig P P Huang J J Kostrzynska M M Trust T J TJ

Journal of bacteriology 19951101 21

The ability of certain strains of Helicobacter pylori to cause sialic acid-sensitive agglutination of erythrocytes has been attributed to the HpaA protein (D.G. Evans, T.K. Karjalainen, D. J. Evans, Jr., D. Y. Graham, and C.H. Lee, J. Bacteriol. 175:674-683, 1993), the gene for which has been cloned and sequenced. On the basis of the hydropathy plot of HpaA and the presence of a potential lipoprotein signal sequence and modification site, and because of the similarities of these features with th ...[more]

PMID: 7592366

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Project description:Our current understanding of lipoprotein synthesis and localization in Gram-negative bacteria is based primarily on studies of Escherichia coli Newly synthesized E. coli prolipoproteins undergo posttranslational modifications catalyzed by three essential enzymes (Lgt, LspA, and Lnt). The mature lipoproteins are then sorted to the inner or outer membrane via the Lol system (LolABCDE). Recent studies suggested that this paradigm may not be universally applicable among different classes of proteobacteria. In this study, we conducted a systematic analysis of lipoprotein processing and sorting in Helicobacter pylori, a member of the Epsilonproteobacteria that colonizes the human stomach. We show that H. pylorilgt, lspA, and lnt homologs can complement conditionally lethal E. coli mutant strains in which expression of these genes is conditionally regulated. Mutagenesis studies and analyses of conditionally lethal H. pylori mutant strains indicate that lgt and lspA are essential for H. pylori growth but lnt is dispensable. H. pylorilolA and the single lolC (or lolE) homolog are also essential genes. We then explored the role of lipoproteins in H. pylori Cag type IV secretion system (Cag T4SS) activity. Comparative analysis of the putative VirB7 homolog CagT in wild-type and lnt mutant H. pylori strains indicates that CagT undergoes amino-terminal modifications consistent with lipidation, and we show that CagT lipidation is essential for CagT stability and Cag T4SS function. This work demonstrates that lipoprotein synthesis and localization in H. pylori diverge from the canonical pathways and that lipidation of a T4SS component is necessary for H. pylori Cag T4SS activity.IMPORTANCE Bacterial lipoproteins have diverse roles in multiple aspects of bacterial physiology, antimicrobial resistance, and pathogenesis. Dedicated pathways direct the posttranslational lipidation and localization of lipoproteins, but there is considerable variation in these pathways among the proteobacteria. In this study, we characterized the proteins responsible for lipoprotein synthesis and localization in Helicobacter pylori, a member of the Epsilonproteobacteria that contributes to stomach cancer pathogenesis. We also provide evidence suggesting that lipidation of CagT, a component of the H. pylori Cag T4SS, is required for delivery of the H. pylori CagA oncoprotein into human gastric cells. Overall, these results constitute the first systematic analysis of H. pylori lipoprotein production and localization pathways and reveal how these processes in H. pylori differ from corresponding pathways in model proteobacteria.

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Dataset Information

The putative neuraminyllactose-binding hemagglutinin HpaA of Helicobacter pylori CCUG 17874 is a lipoprotein.

Publications

The putative neuraminyllactose-binding hemagglutinin HpaA of Helicobacter pylori CCUG 17874 is a lipoprotein.

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