Our goal has been to determine sensitive targets in the HIV-1 protease and then develop specific inhibitors of the enzyme. We are using the knowledge that metals, such as copper, are inhibitors of the protease to locate these sites. Mutant forms of the protease were used in order to study copper inhibitors of the protease. We found that inhibition of the HIV-1 protease by copper (I) and copper (II) is dependent on the cysteines in the protease and that each cysteine is involved in copper mediated inhibition. Copper complexes were also found to be good inhibitors of the protease. Inhibition by an organic copper (I) complex was not dependent on the cysteines in the protease while inhibition by a penicillamine copper complex was cysteine dependent. Moreover, an organic copper (I) complex was found to inhibit the replication of HIV-1 in cells. To determine which of the cysteines of the protease could be a target for HIV-1 protease inhibition we modified the cysteines and assessed enzyme function. It was found that modifying cysteine-95 completely inhibited enzyme activity while modifying cysteine-67 reduced activity by about 50%. This inhibition was reversible when the label was removed with dithiothreitol. The HIV-1 protease contains two reactive cysteines which are highly conserved. They may play a regulatory role in HIV-1 maturation and they provide a possible target for inhibition of the protease.