FIG. 6.

Interactions of VacA-(Δ6–27), s1/m1 VacA, and chimeric s2/m1 VacA with c-myc-tagged s1/m1 VacA. Three species of purified VacA were biotinylated as described in Materials and Methods. Samples used for immunoprecipitations included acid-activated biotinylated VacA (lane a), acid-activated mixtures of biotinylated VacA and c-myc-tagged VacA (lane b), and untreated mixtures of biotinylated VacA and c-myc-tagged VacA (lane c). Immunoprecipitations using anti-c-myc antibody 9E10 were performed as described in Materials and Methods. Immunoprecipitated proteins were separated by SDS-PAGE and transferred to nitrocellulose, and biotinylated VacA was detected with streptavidin-conjugated horseradish peroxidase and enhanced chemiluminescence. Biotinylated VacA species were successfully immunoprecipitated only if mixtures of biotinylated VacA and c-myc VacA were acid activated prior to the immunoprecipitation (lane b).