Figure 2. Expression and function of single Trp mutants in yeast.

A, 15 µg of crude microsomal membrane protein from yeast expressing Wt Pgp, empty vector (negative control), and each Trp mutant was analyzed by Western blot using the C219 anti-Pgp antibody. A representative of at least three experiments is shown; protein MW marker positions are indicated in kDa. B, Fungicidal resistance was measured by growing mutant expressing yeast and control strains in the absence or presence of 50 µM FK506 and measuring the OD₆₀₀ at 24 to 25 hours. Relative growth is the OD₆₀₀ of cells grown in drug/the OD₆₀₀ of cells grown in drug-free medium. Bars represent the mean relative growth ± SEM of triplicate samples from three or more independent experiments normalized to the relative growth of yeast expressing Wt Pgp. C, Mating efficiency of Trp mutants was determined by mating the a-type Trp mutant expressing cells with an α-type tester strain and plating on minimal media selective for diploids. Relative mating efficiency is the number of diploid cells divided by the total number of cells, then divided by the mating efficiency of Wt expressing yeast. Bars indicate the mean relative drug resistance ± SEM from four independent experiments. (*) indicate samples that were significantly different than Wt using a two tail t-test with p≤0.02.