The Apicomplexan Molecular Physiology Unit conducts basic research on the transport of ions and nutrients across various membranes of human red blood cells infected with malaria parasites. This work incorporates molecular biology and informatics, protein and lipid biochemistry, immunofluorescent localization of membrane proteins, various transport assays, biophysics, high-throughput screening of compound libraries, and examination of structure-activity relationships for small molecule inhibitors. We previously identified an unusual ion channel on human red blood cells infected with P. falciparum, which causes the deadliest form of malaria. This channel, the plasmodial surface anion channel (PSAC), is present at 1000 copies/cell, has unusual gating properties, and is permeable to a range of anions and nutrients known to be required for parasite growth. We proposed that PSAC mediates the first step in a sequential diffusive pathway of nutrient acquisition. Current projects in the lab include: 1) functional studies to examine PSAC gating and selectivity properties, 2) high-throughput screening to identify high affinity, high specificity PSAC antagonists with therapeutic potential, 3) biochemical and molecular biological studies aimed at cloning the gene(s) encoding PSAC and other transporters, and 4) heterologous expression of these transporters. Our overall goal with these projects is to probe how PSAC achieves its unusual functional properties, to understand the parasite's cell biology and physiology, and to develop new strategies for the control of malaria. In the past fiscal year, the lab made several contributions to this important field. Most importantly, we examined a fundamental debate on whether PSAC is a parasite-encoded protein trafficked to the host RBC membrane or a modified host membrane protein. In these studies, we identified, for the first time, polymorphisms in PSAC gating behavior when red cells are cultured with geographically distinct parasite isolates, suggesting that PSAC is parasite-encoded. A second accomplishment is the identification of dantrolene as an unusual PSAC antagonist, providing an explanation for its in vitro parasite growth inhibitory effects. A secondary screen of 164 dantrolene derivatives then identified compounds of higher specificity and affinity than any previously known PSAC antagonists. Finally, we probed channel gating with and without added antagonists to determine that PSAC has only one open and at least three closed states. Our studies have provided new insights into this unusual channel's structure and function.
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