The overall objective of this proposal is to understand the structure, function, and developmental regulation of the mitochondrial ATP synthase of the parasitic protozoan Trypanosoma brucei. This critical enzyme couples membrane potential generated by the electron transport chain to synthesis of ATP. We have recently established that the enzyme is regulated through the life cycle of T. brucei, although not to the same extent as the cytochromes of the electron transport chain. We now wish to determine the nature of regulation of the ATP synthase through the bloodstream of the mammalian host and in the insect host.
Specific aims are to: 1. Examine the complete structure of the ATP synthase to determine whether the substructure of the complex may vary or whether the protein assembly or stability might play a regulatory role. 2. Establish whether interaction of regulatory proteins with the enzyme complex may be a component of the regulation of the ATP synthase in the developmental cycle. 3. Determine the primary sequence of genes encoding critical subunits of the ATP synthase at the genomic and cDNA level, to establish whether transcript editing occurs as a function of development and to determine whether editing occurs with both nuclear and mitochondrially encoded proteins. 4. Establish whether regulation of the ATP synthase complex may occur at the level of protein expression in T. brucei. Results from these experiments will give a better understanding of the structure and functional status of the ATP synthase and may yield insights into its role in energy metabolism through the developmental stages of T. brucei. We hope that the information we obtain will enable us to specifically target this bioenergetic enzyme complex by chemical means. Due to its central role in energy metabolism we believe it is an excellent candidate for such a chemotherapeutic approach.
