FIG. 6.

Mutational analysis of VacA residues 346 and 347. Full-length WT VacA and a panel of VacA proteins containing mutations in residues 346 and/or 347 were expressed in E. coli. E. coli soluble extracts containing the indicated recombinant proteins were normalized based on immunoblotting so that they contained equivalent concentrations of VacA and were then added to the medium overlying HeLa cells. Vacuolating activity was quantified by using a neutral red uptake assay. The results represent the mean ± the SD from triplicate samples. *, P ≤ 0.05 as determined by using ANOVA, followed by Dunnett's post hoc test compared to WT VacA.