ISOLATION AND CHARACTERIZATION OF 2 CAMPYLOBACTER GLYCINE-EXTRACTED PROTEINS THAT BIND TO HELA-CELL MEMBRANES

Two immunogenic proteins of 27 (CBF1) and 29 (CBF2) kDa from enteropat hogenic Campylobacter species appear to bind to mammalian cells. We pu rified these two proteins from a pathogenic and adherent Campylobacter jejuni strain to homogeneity by using acid extraction, preparative ge l electrophoresis, and electroelution. Polyclonal rabbit antisera to t hese proteins were prepared. Immunologic studies indicate that CBF1 co rresponds to the PEB1 and CBF2 corresponds to the PEB4 described by Pe i et al. (Z. Pei, R. T. Ellison, and M. Blaser, J. Biol. Chem. 226:163 63-16369, 1991). Immunogold labeling of a C. jejuni adherent strain wi th anti-CBF1, anti-CBF2, and anti-PEB1 suggested that CBF1 (PEB1) is s urface exposed while CBF2 (PEB4) is not. Analysis of whole-cell extrac ts from 14 strains by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis with 7 M urea and immunoblotting with antisera to CBF1 and CBF2 suggests that CBF proteins from adherent and nonadherent str ains are different. Use of purified proteins in a microassay of adhere nce to cellular membranes indicated that CBF1 was much more adherent t han CBF2. Adherence of C. jejuni to viable HeLa cells was markedly red uced with the antiserum to CBF1, whereas the CBF2 antiserum was a poor inhibitor. Purified CBF1 competitively inhibited adherence of whole b acteria to HELa cells, whereas purified CBF2 was no better a competito r than bovine serum albumin. Adherence of CBF2 was markedly reduced in the presence of Tween 20 or SDS, whereas adherence of CBF1 was reduce d only by SDS. We conclude that (i) CBF1 (PEB1) is surface exposed and may be the key protein for C. jejuni adhesion and (ii) CBF2 (PEB4) ma y be complexed with CBF1 and may passively coadhere with CBF1 under ce rtain experimental conditions. Adherent and nonadherent strains contai n different isotypes of these two proteins which could be useful marke rs of C. jejuni adhesion.