Two-dimensional gel electrophoresis and immunoblotting of Campylobacter outer membrane proteins

Abstract

We characterized outer membrane proteins (OMPs) from selected Campylobacter jejuni, C. coli, and C. fetus strains by two-dimensional gel electrophoresis (2DGE), using isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and by immunoblotting with immune rabbit serum. The flagellar band with a molecular mass of 63 kilodaltons (kDa) demonstrated previously by one-dimensional SDS-PAGE was shown by 2DGE to consist of one or several charge trains, depending upon the species, strain, and type of preparation studied; each of the individual peptides was found to be antigenic by immunoblotting. In contrast, in all of the strains studied, the major OMP (43 to 44 kDa) of C.jejuni and C. coli consisted of a single isomeric form which was weakly immunogenic. Several minor proteins (29 to 31 kDa) were found to be strongly immunogenic by immunoblotting. C. fetus strains possessed two major OMPs of 45 to 47 kDa, each of which consisted of either a single isomer or a major isomer comprising at least 90% of the major OMP. Serum-resistant strains of C. fetus possessed an acid-labile 100-kDa glycoprotein (pI, 4.1) which was markedly diminished or absent in serum-sensitive strains. These 2DGE analyses provide information that is useful in taxonomic and epidemiologic studies and for the purification of surface antigens for the development of campylobacter vaccines and may also facilitate the identification of specific virulence factors.