Cryptosporidium parvum causes one of the opportunistic infections (OI) in AIDS patients for which no treatment is yet known. Although many drugs have been tested against the OI, knowledge about the presence of target enzymes, biosynthetic pathways, and expression or regulation are virtually unknown. The applicant's preliminary data indicate that several fatty acid synthase (FAS) genes are present in C. parvum (CpFAS) and that their size, organization and functional domains differ from those of humans. These genes appear to be functional, since their transcripts were detected in sporozoites and intracellular stages and the growth of C. parvum in vitro was inhibited by a FAS inhibitor. The discovery of CpFAS genes suggests that this apicomplexan might synthesize fatty acids de novo, thus providing clues for drug development against cyptosporidiosis. Because most parasitic protozoa are auxotrophs for fatty acids, this discovery also opens a new vista for protozoan biochemistry. The investigator's long term goal is to study the potential of apicomplexan fatty acid biosynthesis as a useful target for drug development.
The specific aims of this proposal are: 1.Characterize the CpFAS gene structure, transcription level and subcellular localization during the life cycle of the parasite. 2. Elucidate the function of CpFAS by the analysis of substrate preference, intermediate and final fatty acid products using recombinant CpFAS proteins expressed in bacteria or yeast 3.Determine whether fatty acid biosynthesis can serve as a rational target for drug development by screening FAS analogues against C. parvum in vitro and in vivo. Completion of these specific aims will either support or refute the investigator's hypothesis that fatty acid biosynthesis in C. parvum is sufficiently different from its host to be exploited for drug design target against cryptosporidiosis. The outcome expected is to overcome a major obstacle for therapy against this OI of AIDS patients.
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