Cloning, nucleotide sequence, and expression of a gene encoding an adhesin subunit protein of Helicobacter pylori

D. G. Evans, T. K. Karjalainen, D. J. Evans, D. Y. Graham, Chao-Hung Lee

Research output: Contribution to journalArticle

162 Citations (Scopus)

Abstract

Gene hpaA, which codes for the receptor-binding subunit of the N- acetylneuraminyllactose-binding fibrillar hemagglutinin (NLBH) of Helicobacter pylori, was cloned and sequenced. The protein expressed by hpaA, designated HpaA, was identified as the adhesin subunit on the basis of its fetuin-binding activity and its reactivity with a polyclonal, monospecific rabbit serum prepared against NLBH purified from H. pylori. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and Western blots (immunoblots) showed that the cloned adhesin has the same molecular weight (20,000) as that found on H. pylori. Also, HpaA contains a short sequence of amino acids (KRTIQK) which are all either identical or functionally similar to those which compose the sialic acid-binding motif of Escherichia coli SfaS, K99, and CFA/I. Affinity-purified antibody specific for a 12-residue synthetic peptide that included this sequence blocked the hemagglutinating activity of H. pylori and was shown by immuno-gold electron microscopy to react with almost transparent material on unstained H. pylori cells, which is consistent with previous observations concerning the location and morphology of the NLBH.

Original languageEnglish (US)
Pages (from-to)674-683
Number of pages10
JournalJournal of Bacteriology
Volume175
Issue number3
StatePublished - 1993
Externally publishedYes

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Protein Subunits
Helicobacter pylori
Organism Cloning
Gene Expression
Western Blotting
Fetuins
Antibody Affinity
Immunoelectron Microscopy
Hemagglutinins
N-Acetylneuraminic Acid
Sodium Dodecyl Sulfate
Gold
Polyacrylamide Gel Electrophoresis
Amino Acid Sequence
Molecular Weight
Escherichia coli
Rabbits
Peptides
Serum
Genes

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Immunology

Cite this

Cloning, nucleotide sequence, and expression of a gene encoding an adhesin subunit protein of Helicobacter pylori. / Evans, D. G.; Karjalainen, T. K.; Evans, D. J.; Graham, D. Y.; Lee, Chao-Hung.

In: Journal of Bacteriology, Vol. 175, No. 3, 1993, p. 674-683.

Research output: Contribution to journalArticle

Evans, D. G. ; Karjalainen, T. K. ; Evans, D. J. ; Graham, D. Y. ; Lee, Chao-Hung. / Cloning, nucleotide sequence, and expression of a gene encoding an adhesin subunit protein of Helicobacter pylori. In: Journal of Bacteriology. 1993 ; Vol. 175, No. 3. pp. 674-683.
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AU - Graham, D. Y.

AU - Lee, Chao-Hung

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N2 - Gene hpaA, which codes for the receptor-binding subunit of the N- acetylneuraminyllactose-binding fibrillar hemagglutinin (NLBH) of Helicobacter pylori, was cloned and sequenced. The protein expressed by hpaA, designated HpaA, was identified as the adhesin subunit on the basis of its fetuin-binding activity and its reactivity with a polyclonal, monospecific rabbit serum prepared against NLBH purified from H. pylori. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and Western blots (immunoblots) showed that the cloned adhesin has the same molecular weight (20,000) as that found on H. pylori. Also, HpaA contains a short sequence of amino acids (KRTIQK) which are all either identical or functionally similar to those which compose the sialic acid-binding motif of Escherichia coli SfaS, K99, and CFA/I. Affinity-purified antibody specific for a 12-residue synthetic peptide that included this sequence blocked the hemagglutinating activity of H. pylori and was shown by immuno-gold electron microscopy to react with almost transparent material on unstained H. pylori cells, which is consistent with previous observations concerning the location and morphology of the NLBH.

AB - Gene hpaA, which codes for the receptor-binding subunit of the N- acetylneuraminyllactose-binding fibrillar hemagglutinin (NLBH) of Helicobacter pylori, was cloned and sequenced. The protein expressed by hpaA, designated HpaA, was identified as the adhesin subunit on the basis of its fetuin-binding activity and its reactivity with a polyclonal, monospecific rabbit serum prepared against NLBH purified from H. pylori. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and Western blots (immunoblots) showed that the cloned adhesin has the same molecular weight (20,000) as that found on H. pylori. Also, HpaA contains a short sequence of amino acids (KRTIQK) which are all either identical or functionally similar to those which compose the sialic acid-binding motif of Escherichia coli SfaS, K99, and CFA/I. Affinity-purified antibody specific for a 12-residue synthetic peptide that included this sequence blocked the hemagglutinating activity of H. pylori and was shown by immuno-gold electron microscopy to react with almost transparent material on unstained H. pylori cells, which is consistent with previous observations concerning the location and morphology of the NLBH.

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