Molecular Screening For Antagonists Of Hiv-1 Viral Infectivity Factor Enables Drug Discovery

HIV-1 Viral infectivity factor (Vif) is a viral accessory protein that is absolutely required for HIV-1 infection in CD4+ T cells, macrophages and dendritic cells due largely to its role in recruiting antiretroviral factors of the APOBEC3 family to an E3 ligase complex for polyubiquitination and proteasomal degradation. Oligomerization of Vif is essential for APOBEC3G degradation and for the HIV-1 life-cycle but the mechanism of action and molecular oligomeric interface are unknown. Vif oligomerization is mediated by a conserved 161PPLP164 motif, but the crystal structure of Vif (PDB 4N9F) revealed the burial of the PPLP motif between N and C-terminal domains. The sole Vif-Vif crystal contact involves the conserved 40YRHHY44 motif, potentially implicating its involvement in a biological Vif oligomeric interface. The YRHHY motif is also essential for A3G degradation. Thus, inhibition of Vif-dependent A3G degradation and HIV infectivity by PPLP peptidomimetics may be allosteric. OyaGen has identified a small molecule Vif dimerization antagonist through a novel in-cell HTS quenched-FRET assay. Here we show that this hit, BRD25, disrupts Vif oligomerization, reduces APOBEC3G degradation, and decreases HIV infectivity. Fluorescence anisotropy was used to show that BRD25 reduces the size of purified Vif in a dose-dependent manner, consistent with oligomer disruption. In silico docking of BRD25 with the crystal structure of Vif reveals a potential binding site consistent with its role in disruption of Vif oligomerization and A3G degradation. These data demonstrate the availability of protein-protein interactions as novel drug targets in HIV infected cells.