The purpose of the proposed research is to utilize the feline lentivirus, FIV, as a system for testing strategies aimed at the rational design of antivirals efficacious against lentivirus infections. The similarities between FIV and HIV in life cycle, genomic make-up, and etiology of disease resulting from infection make the cat a particularly relevant model. In addition, the relative economy, safety, and manipulability of the feline system make it much more amenable to detailed testing than any of the primate systems. Studies will center around the characterization of FIV protease (PR) and the sites within FIV that are cleaved by this critical enzyme. Both eucaryotic and procaryotic expression clones will be prepared in order to 1) determine PR cleavage sites; 2) establish a hierarchy of cleavage; 3) assess cleavage rates of the natural and synthetic PRs against native substrates; and 4) monitor the influence of drugs arising from Projects 1-4 on Pr cleavage of natural substrates. The data generated will be used for synthetic substrate preparation in Project 4; drug design by projects 1, 2 and 3; for comparison of cleavage specificities with HIV PR; and to aid in the establishment of consensus rules for retroviral PR cleavage. This project will also be responsible for the preparation of site-directed mutants of PR, based on predicted interactions between enzyme and drug, established in Projects 1-4. We will also carry out all virological aspects of the program, which will involve the in vitro testing of selected drugs for both toxicity and antiviral efficacy. The results of these studies will fulfill the anchor phase of the developmental loop, providing support and feedback for design strategies arising from the overall program.