The 3-O-(3’,3’-dimethylsuccinyl) derivative of betulinic acid (DSB) inhibits the assembly of virus-like particles in HIV-1 Gag precursor-expressing cells

Abstract

Background The 3- O-(3’,3’-dimethylsuccinyl) derivative of betulinic acid (DSB) blocks HIV-1 maturation by interfering with viral protease (PR) at the capsid (CA)-SP1 cleavage site, a crucial region in HIV-1 morphogenesis. Methods We analysed the effect of DSB on the assembly of HIV-1 Gag precursor (Pr55GagHIV) into membrane-enveloped virus-like particles (VLP) in baculovirus-infected cells expressing Pr55GagHIV, in a cellular context devoid of viral PR. Results DSB showed a dose-dependent negative effect on VLP assembly, with an IC50∼10 μM. The DSB inhibitory effect was p6-independent and was also observed for intracellular assembly of non-N-myristoylated Gag core-like particles. HIV-1 VLP assembled in the presence of DSB exhibited a lower stability of their inner cores upon membrane delipidation compared with control VLP, suggesting weaker Gag-Gag interactions. DSB also inhibited the assembly of simian immunodeficiency virus SIVmac251 VLP, although with a twofold lower efficacy (IC50∼20 μM). No detectable inhibitory activity was observed for murine leukaemia virus (MLV) VLP; however, fusion of the SP1-NC-p6 domains from HIV-1 to the matrix (MA)-CA domains from MLV conferred DSB sensitivity to the chimaeric Gag precursor Pr72GagMLV–HIV (IC50=30 μM). This observation suggested that the main DSB target on Pr55Gag was the SP1 domain, but the higher degree of DSB resistance for Pr72GagMLV–HIV compared with Pr55GagHIV implied that other upstream Gag region(s) might contribute to DSB reactivity. Conclusions Sequence alignment and three-dimensional modelling by homology of the CA-SP1-NC junction in HIV-1, SIVmac251 and Pr72GagMLV–HIV suggested that a higher hydrophilic character of the CA region immediately upstream to the HIV-1 CA-SP1 junction, as occurred in Pr72GagMLV–HIV, correlated with a lower DSB sensitivity.