Trichomoniasis is the most common non-viral sexually transmitted disease world-wide and is caused by the human parasite Trichomonas vaginalis. Currently only two drugs are approved for treatment of trichomoniasis, both of which are 5-nitroimidazoles that appear to use the same mechanisms to kill the parasite, and share a site of action, the hydrogenosome, a unique energy-producing organelle. The goal of the proposed studies is to elucidate the identity and function of a potential inner membrane translocase of the hydrogenosome in T. vaginalis. One member of the translocase complex, Hmp24, has been identified and knocked out in these cells. The AHmp24 cells exhibit a decrease in the kinetics of import into the hydrogenosome, confirming that Hmp24 does indeed play a role in protein import. The proposed experiments would further define the role of Hmp24 in import by examining the expression pattern of other members of the Hmp24 gene family and creation of a double-knockout with one of these paralogs, Hmp24a. Identification of other members of the Hmp24 protein complex will be done by precipitation of the complexes using tagged Hmp24 followed by mass spectrometry. Formation of import-competent complexes will also be examined in AHmp24/24a cell lines. The role of specific domains of Hmp24 on both complex formation and import into the hydrogenosome will be examined through mutagenesis experiments. The findings of these studies will illuminate possible new drug targets for trichomoniasis as well as elucidate further the evolutionary relationship between the hydrogenosome and the mitochondrion of other eukaryotes. PUBLIC HEALTH RELEVANCE: Because of increased resistance to treatment and the important health consequences of infection, namely increased susceptibility to HIV infection and cervical cancer in women, new research is needed to identify new drug targets within the parasite. The hydrogenosome is an appealing target for any new drug, both because it is the energy-production site within the parasite and because it presents a unique target not present in host cells.