THE CENTRAL VARIABLE V2 REGION OF THE CS31A MAJOR SUBUNIT IS INVOLVEDIN THE RECEPTOR-BINDING DOMAIN

CS31A is a K88-related capsule-like surface protein that mediates Esch erichia coli and Klebsiella pneumoniae adhesion to the human Caco-2 an d Intestine-407 cell lines, In this study, we demonstrate that ClpG, t he major subunit of CS31A, contains the adhesive domain of the polymer ized structure, We mapped this domain within the ClpG protein by perfo rming adhesion inhibition experiments with Intestine-407 cells with ni ne synthetic peptides (CLP1 to CLP9) covering the dominant antigenic r egions of ClpG and with the corresponding rabbit antipeptide antibodie s. The peptides CLP1 (amino acid positions in parentheses) (5-18), CLP 2 (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) p eptide, corresponding to the central part of the protein, and relevant antibodies inhibited bacterial adhesion to intestinal epithelial cell s, Anti-CLP4 (97-109) and anti-CLP6 (148-162) antibodies targeting reg ions surrounding the CLP5 sequence also inhibited bacterial adhesion, Site-directed mutagenesis experiments inducing changes in the amino ac id sequence of the ClpG protein corresponding to the CLP5 peptide resu lted in the expression of nonadhesive CS31A antigen. These findings in dicate that the ClpG receptor-binding domain is located in the central variable V2 region.