FIG. 5.

Subcellular localization of SpiC-M45. Wild-type Salmonella serovar Typhimurium strain NCTC 12023 carrying p2127 was grown under conditions inducing the expression and secretion of SPI2 substrate proteins. Bacteria were harvested and subjected to subcellular fractionation as described in Materials and Methods. The various fractions (M, total membrane fraction; S, soluble fraction; I, inner membrane fraction; O, outer membrane fraction) were subjected to SDS-PAGE on 12% Tricine gels; protein was transferred onto nitrocellulose membranes, and the M45 epitope was detected with monoclonal antibodies and an ECL kit (Amersham). The purity of the various fractions was confirmed by Western blotting using antibodies against subunit β of ATP synthase and OmpA as tracers for the inner membrane and outer membrane fractions, respectively.