DNA BASE EXCISION REPAIR IN PLASMODIUM FALCIPARUM

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DESCRIPTION: (provided by the applicant): DNA replication and repair, processes that are essential for cellular proliferation and the maintenance of genome integrity, have gone largely unexplored in parasitology. The replication and control of DNA repair in Plasmodium falciparum are likely to have a number of differences from the mammalian cell. The rapid replication as trophozoites mature into schizonts and the three rounds of replication that microgametes accomplish in 10 minutes at exflagellation are both unusual. For this reason, a basic understanding of these processes in the parasite is an important goal. Our groundbreaking investigations revealed that Plasmodium falciparum (Pj) has DNA repair capabilities, and that the removal of uracil and the repair of abasic sites in DNA are exclusively by a long-patch base excision repair (BER) pathway, which is remarkably different from mammalian cells. Important quantitative differences between the enzymology of the BER pathway in P. falciparum and mammalian cells have been established. The short-term goal of this proposal is to define the fundamental reaction of BER in Plasmodium falciparum. The essential proteins in the parasite BER pathway have been identified and the over expression of the enzymes in bacteria will facilitate their biochemical characterization. These enzymes include Pf AP endonuclease, Pf flap endonuclease (Pf FEN-1) and Pf DNA ligase. Expression of these enzymes is part of our plan to reconstitute the entire parasite BER pathway for studying the individual contributions of each of the key enzymes to the long-patch repair process. A reconstituted BER system will afford future study of the various protein-protein interactions, mechanisms, associations and biochemical reactions under controlled situations of BER. The ultimate goal of these studies is to identify the Plasmodium excision repair genes, determine their biological and biochemical function and assess their role in maintaining the parasite genome. Completion of the proposed aims in this project will provide important, new information about a neglected area of parasite biology, and possibly the opportunity to exploit differences in DNA repair between P. falciparum and mammalian cells to develop new strategies for antimalarial therapy.