Targeting Trypanosoma brucei S-adenosylmethionine decarboxylase in Drug Discovery

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DESCRIPTION (provided by applicant): Human African Trypanosomiasis (HAT), commonly called sleeping sickness, is caused by single-celled parasites, Trypanosoma brucei, which are transmitted to humans by infected tsetse flies. Trypanosomiasis has a profound impact on the health of a large number of people in sub-Saharan Africa. It is fatal if left untreated and current drug therapy is problematic because of toxicity and emerging resistance. The polyamine biosynthetic pathway is a validated target for the development of drugs against HAT and the emerging evidence provides a compelling case that S-adenosylmethionine decarboxylase (AdoMetDC) is a highly promising target for the development of new anti-trypanosomal agents. AdoMetDC catalyses the ratelimiting step in the production of the polyamine spermidine from putrescine. Mammalian AdoMetDC is a homodimer that uses a pyruvoyl group as a cofactor to stabilize the carbanion intermediate formed during the decarboxylation of AdoMet. Recently, our lab demonstrated that the trypanosomatid AdoMetDC is regulated by a unique mechanism, heterodimer formation with a catalytically dead AdoMetDC homolog. This protein, designated prozyme, forms a high-affinity heterodimer with AdoMetDC and increases its activity by >1,000-fold. Further we confirmed that the heterodimer is the functional enzyme in vivo. My primary goals are to elucidate the mechanisms that regulate the AdoMetDC activation by the prozyme and to identify and characterize novel AdoMetDC inhibitors essential for the development of new anti-trypanosomal drugs. In order to do that, I will first focus on the identification of crucial amino acid residues involved in the activation in an attempt to gain insights into the mechanism of allosteric regulation. Next I will characterize species differences in the activation of AdoMetDC by prozyme by comparing the T. brucei AdoMetDC/prozyme complex to the homologs from T. cruzi and L. donovani. Finally I will perform a high-throughput screen (HTS) of a small molecule compound library to identify novel inhibitors of T. brucei AdoMetDC and to test these for activity against cultured parasites. Upon completion of these studies I will have identified and validated a number of novel inhibitors of AdoMetDC for future lead optimization studies that may lead to the discovery of new, less toxic drugs for the treatment of HAT. Project Narrative: Human African Trypanosomiasis is a insect borne disease that is fatal if untreated, yet current drug therapies are toxic and difficult to administer. The work described in this proposal characterizes an essential enzyme that has unique features in the parasite, with the goal of developing new chemotherapeutic agents against this devastating disease.