FIG. 5.
Western blot analysis of invB-dependent expression and secretion of SopEM45 and SipAM45. (A) SL1344 (wild type), SB161 (ΔinvG), and M568 (invB::aphT) carrying the sopEM45 expression plasmid pM136 were grown under SPI-1-inducing conditions (see Materials and Methods). One hundred microliters of a whole culture and 100 μl of a culture supernatant were separated by SDS-PAGE and analyzed by Western blotting by using an anti-M45 (α-M45) antibody. α-OmpC, anti-OmpC antibody. (B) SL1344 (wild type), SB566 (invC::aphT), and M568 (invB::aphT) carrying the sipAM45 expression plasmid pM580, as well as M568 harboring invB (pM249) and sipAM45 (pM580) expression plasmids, were grown under SPI-1-inducing conditions (see Materials and Methods). Twenty-five microliters of a whole culture, pelleted bacteria from 25 μl of a culture, and proteins recovered from 300 μl of a culture supernatant (and dilutions for SL1344) were analyzed by Western blotting by using an anti-M45 (α-M45) antibody. The blots were reprobed by using an anti-OmpC (α-OmpC) antibody to control the amount of protein loaded.