VIEW ARTICLE

Physiology and Biochemistry

Immunochemical Characterization of a Subspecies-Specific Antigenic Determinant of a Membrane Protein Extract of Xanthomonas campestris pv. campestris. N. Thaveechai, Department of Plant, Soil and Entomological Sciences, University of Idaho, Moscow 83843, Present address: Department of Plant Pathology, Kasetsart University, Bangkok 10903, Thailand; N. W. Schaad, Department of Plant, Soil and Entomological Sciences, University of Idaho, Moscow 83843. Phytopathology 76:148-153. Accepted for publication 23 August 1985. Copyright 1986 The American Phytopathological Society. DOI: 10.1094/Phyto-76-148.

A subspecies-specific antigenic determinant(s) of membrane proteins of Xanthomonas campestris pv. campestris was identified and characterized. Extracts of membrane proteins contained sugar, protein, and lipopolysaccharide (LPS) with a relative molecular mass >2,000 kilodaltons (kDas). Treatments of membrane proteins with protease Type VII and XIV, trypsin, lysozyme, beta-galactosidase nucleases, periodate, acid, alkali, detergents, or heat at 100 C for 30 min failed to destroy the immunogen responsible for the specific line of precipitin but those treatments did result in additional lines in Ouchterlony double diffusion and immunoelectrophoresis tests. On the other hand, protease Type 1 destroyed the immunogen. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of membrane proteins showed major peptide bands of 125, 100, 44, 34, 29, and 23.4 kDas. Crossed immunoelectrophoresis (two-dimensional) of SDS-PAGE gels revealed that the specific antigenic determinant was present in the first three bands, the uppermost part of stacking gel, and at the interface between the stacking and resolving gels. Antiserum to purified 125-, 100-, and 44-kDa peptides resulted in reactions of identity when tested against membrane proteins or purified homologous peptides of membrane proteins in Ouchterlony double diffusion tests. Those peptide bands (visualized with SDS-PAGE) with the specific antigenic determinant contained sugar and protein but no LPS. The specific antigenic moiety of membrane proteins was identified as a heat-stable inner membrane peptide.