The HIV-1 protease is essential for viral replication and has provided a very effective target for antiviral drugs to treat AIDS patients. However, the long-term effectiveness of current antiviral protease inhibitors is limited by the rapid development of resistant protease mutants. It is necessary to understand the molecular basis for the action of HIV protease and its inhibitor-resistant mutants in order to develop new inhibitors and new therapeutic strategies. Analysis of crystal structures and substrate specificity has been used to identify important inhibitor-protease interactions and the protease residues that are critical for recognition of substrates. These key residues were predicted to occur in the inhibitor resistant variants of HIV protease and are among the most commonly mutated residues in the known resistant variants. The structures and activities of HIV-1 protease mutants found in resistant isolates have been characterized. Analyses by the PI's group have demonstrated that resistant mutants have altered catalytic activity and specificity for the cleavage of peptides representing critical steps in polyprotein processing, and altered dimer stability. All these effects are expected to contribute to increased viral replication in the presence of protease inhibitors. Computational methods have been developed to predict the relative efficiency of cleavage of peptide substrates of HIV protease, and relative inhibition of protease mutants. Clinical inhibitors will be compared to define the factors important for improved inhibition of resistant protease variants. Structure-based designs will assist with development of new protease inhibitors as the next generation of antiviral agents to overcome resistance.