9509144 Jonsson This research addresses the biochemistry and genetics of the human T-cell leukemia virus (HTLV) integration pathway. Specifically, the underlying basis for integrase (IN)-LTR recognition will be defined. One approach will be to construct chimeric HTLV-I and HTLV-II integrases to identify the LTR recognition domain. Import of the amino acid domain for recognition of HTLV-I IN's LTR to HTLV-IIg IN would correspond to a loss of recognition for the HTLV-IIg LTR and gain of recognition for the HTLV-I LTR by the HTLV-II/I chimeric protein. Once the domain has been identified, site-directed mutagenesis will be employed to ascertain the important functional amino acid residues. A second aspect of the research will focus on the molecular details of binding and recognition of the LTR by IN using fluorescence technology. This fluorescence approach is useful in that it offers a more dynamic and sensitive approach to binding, which will be important in the light of the "weak" DNA binding activity of integrases. Direct impact of these studies will be toward increased understanding of the biochemical pathway of retroviral integration. In a broader sense, however, the reactions catalyzed by IN have implications in several other areas such as "weak" protein-DNA interactions, and recombinases where scientific advancement has not been as rapid as with other DNA-protein interactions. %%% This research focuses on the protein-DNA interactions of the retroviral protein, integrase. Retroviruses are a large family of single stranded RNA viruses. All retroviruses contain the gene that encodes for integrase, which functions to insert the viral DNA into the cell's DNA. The insertion of the viral DNA is essential to the viral life cycle. In order for integrase to insert or integrate the viral DNA, the protein must "recognize" the viral DNA and the cell's DNA. The specific aim of this project is to define the manner whereby integrase recognizes the retroviral DNA and the cell's DNA. ***
