The central variable V2 region of the CS31A major subunit is involved in the receptor-binding domain.
CS31A is a K88-related capsule-like surface protein that mediates Escherichia coli and Klebsiella pneumoniae adhesion to the human Caco-2 and Intestine-407 cell lines. In this study, we demonstrate that ClpG, the major subunit of CS31A, contains the adhesive domain of the polymerized structure. We mapped this domain within the ClpG protein by performing adhesion inhibition experiments with Intestine-407 cells with nine synthetic peptides (CLP1 to CLP9) covering the dominant antigenic regions of ClpG and with the corresponding rabbit antipeptide antibodies. The peptides CLP1 (amino acid positions in parentheses) (5-18), CLP2 (44-56), CLP3 (82-96), CLP7 (174-190), CLP8 (185-199), and CLP9 (235-249) and corresponding antipeptide antibodies targeting parts of the N- and C-terminal regions of ClpG had no effect on the adhesion of the TCFF15 recombinant strain expressing CS31A. Only the CLP5 (132-146) peptide, corresponding to the central part of the protein, and relevant antibodies inhibited bacterial adhesion to intestinal epithelial cells. Anti-CLP4 (97-109) and anti-CLP6 (148-162) antibodies targeting regions surrounding the CLP5 sequence also inhibited bacterial adhesion. Site-directed mutagenesis experiments inducing changes in the amino acid sequence of the ClpG protein corresponding to the CLP5 peptide resulted in the expression of nonadhesive CS31A antigen. These findings indicate that the ClpG receptor-binding domain is located in the central variable V2 region.
Di Martino
P
P
Laboratoire de Bactériologie, Faculté de Pharmacie, Université d'Auvergne, Clermont-Ferrand, France.
Girardeau
J P
JP
Der Vartanian
M
M
Joly
B
B
Darfeuille-Michaud
A
A
D018830:Adhesins, Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D000595:Amino Acid Sequence; D000907:Antibodies, Bacterial / Q000737:chemistry; D000942:Antigens, Bacterial / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D001422:Bacterial Adhesion / Q000187:drug effects / Q000276:immunology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D002460:Cell Line; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000276:immunology / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis
D018830:Adhesins, Escherichia coli; D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D007074:Immunoglobulin G; D010446:Peptide Fragments; D011994:Recombinant Proteins; C071730:clpG protein, E coli