Abstract
Processing of the Gag precursor protein by the viral protease during particle release triggers virion maturation, an essential step in the virus replication cycle. The first-in-class HIV-1 maturation inhibitor dimethylsuccinyl betulinic acid [PA-457 or bevirimat (BVM)] blocks HIV-1 maturation by inhibiting the cleavage of the capsid-spacer peptide 1 (CA-SP1) intermediate to mature CA. A structurally distinct molecule, PF-46396, was recently reported to have a similar mode of action to that of BVM. Because of the structural dissimilarity between BVM and PF-46396, we hypothesized that the two compounds might interact differentially with the putative maturation inhibitor-binding pocket in Gag. To test this hypothesis, PF-46396 resistance was selected for in vitro. Resistance mutations were identified in three regions of Gag: around the CA-SP1 cleavage site where BVM resistance maps, at CA amino acid 201, and in the CA major homology region (MHR). The MHR mutants are profoundly PF-46396-dependent in Gag assembly and release and virus replication. The severe defect exhibited by the inhibitor-dependent MHR mutants in the absence of the compound is also corrected by a second-site compensatory change far downstream in SP1, suggesting structural and functional cross-talk between the HIV-1 CA MHR and SP1. When PF-46396 and BVM were both present in infected cells they exhibited mutually antagonistic behavior. Together, these results identify Gag residues that line the maturation inhibitor-binding pocket and suggest that BVM and PF-46396 interact differentially with this putative pocket. These findings provide novel insights into the structure-function relationship between the CA MHR and SP1, two domains of Gag that are critical to both assembly and maturation. The highly conserved nature of the MHR across all orthoretroviridae suggests that these findings will be broadly relevant to retroviral assembly. Finally, the results presented here provide a framework for increased structural understanding of HIV-1 maturation inhibitor activity.
Highlights
Over 20 antiretroviral inhibitors have been approved for clinical use in HIV-1-infected patients
Maturation inhibitors prevent the viral protease from processing Gag at one particular cleavage site, but how they accomplish this is not understood
We found that one of these compounds, PF-46396, gives rise to resistance mutations that map to three domains in Gag, including a region known as the major homology region (MHR)
Summary
Over 20 antiretroviral inhibitors have been approved for clinical use in HIV-1-infected patients These drugs fall into several classes, mostly targeting the viral enzymes reverse transcriptase (RT), protease (PR), and integrase (IN). A fusion inhibitor specific for the viral transmembrane envelope (Env) glycoprotein gp, and an entry inhibitor directed against the viral coreceptor CCR5 are available These antiretrovirals, administered in combinations referred to as highly active antiretroviral therapy (HAART), are quite effective and have resulted in striking declines in AIDS-related mortality in treated patients [1,2,3,4]. Resistance to these compounds, as well as a variety of drug tolerability and related compliance issues, have reduced their benefit in many patients. In addition to the potential therapeutic benefit of such new antiretrovirals, drug discovery efforts are likely to provide novel and important insights into the molecular biology of HIV-1 replication
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