Fig 6.

The V656A mutant exhibits WT drug-binding behavior. We carried out photoaffinity labeling with IAAP in the presence and absence of clo as described in Materials and Methods. (A) Each lane contains 10.8 μg of solubilized PM vesicle protein. After electrophoresis, the gels were dried and exposed to an X-ray film. The radioactivity incorporated into the pdr5 band was quantified using a phosphorimager and analyzed using GraphPad Prism. The autoradiogram described above shows IAAP binding to the WT pdr5 and the mutant pdr5 in the presence and absence of clo. The isogenic strain lacking Pdr5 (_pdr5) was used as a control in the experiment. WT and V656A mutant strains were analyzed in the same experiments. (B) Plots of the experiment shown in panel A. ■, WT; ▲, V656A mutant. We performed this experiment an additional time with nearly identical results.