Fig. 4

V5 western blot quantifying relative PfCRT expression in total yeast membranes (CM) vs isolated yeast vacuole membranes (Vac) for unmodified and and PMA-fused PfCRTs. All lanes contain 3 µg total protein. The blot shows that the PMA leader sequence substantially reduces PfCRT expression at the yeast vacuolar membrane relative to unmodified PfCRT (compare unmodified HB3 PfCRT, left two lanes, vs PMA-HB3 PfCRT chimera, middle two lanes). Note that we are unable to isolate purified vacuolar membranes for yeast expressing unmodified PH1 or PH2 PfCRT, since the intrinsic toxicity conferred by these PfCRTs leads to severe growth defects accompanied by serious perturbation of vacuolar function (and presumably vacuolar integrity) over longer periods of time in culture (see text)