Laboratory of Structural Biology Research, National Institute of Arthritis and Musculoskeletal and Skin Diseases, NIH, Building 50, Room 1517, 50 South Drive, MSC 8025, Bethesda, MD 20892, USA.
J Virol. 2011 Feb;85(4):1420-8. doi: 10.1128/JVI.01926-10. Epub 2010 Nov 24.
Maturation of nascent virions, a key step in retroviral replication, involves cleavage of the Gag polyprotein by the viral protease into its matrix (MA), capsid (CA), and nucleocapsid (NC) components and their subsequent reorganization. Bevirimat (BVM) defines a new class of antiviral drugs termed maturation inhibitors. BVM acts by blocking the final cleavage event in Gag processing, the separation of CA from its C-terminal spacer peptide 1 (SP1). Prior evidence suggests that BVM binds to Gag assembled in immature virions, preventing the protease from accessing the CA-SP1 cleavage site. To investigate this hypothesis, we used cryo-electron tomography to examine the structures of (noninfectious) HIV-1 viral particles isolated from BVM-treated cells. We find that these particles contain an incomplete shell of density underlying the viral envelope, with a hexagonal honeycomb structure similar to the Gag lattice of immature HIV but lacking the innermost, NC-related, layer. We conclude that the shell represents a remnant of the immature Gag lattice that has been processed, except at the CA-SP1 sites, but has remained largely intact. We also compared BVM-treated particles with virions formed by the mutant CA5, in which cleavage between CA and SP1 is also blocked. Here, we find a thinner CA-related shell with no visible evidence of honeycomb organization, indicative of an altered conformation and further suggesting that binding of BVM stabilizes the immature lattice. In both cases, the observed failure to assemble mature capsids correlates with the loss of infectivity.
成熟的新生病毒粒子,逆转录病毒复制的一个关键步骤,涉及到病毒蛋白酶将 Gag 多蛋白切割成基质(MA)、衣壳(CA)和核衣壳(NC)成分,以及它们随后的重组。贝伐单抗(BVM)定义了一类新的抗病毒药物,称为成熟抑制剂。BVM 通过阻断 Gag 加工中的最后一个切割事件,即 CA 与其 C 末端间隔肽 1(SP1)的分离,发挥作用。先前的证据表明,BVM 结合在不成熟病毒粒子中组装的 Gag,阻止蛋白酶接近 CA-SP1 切割位点。为了研究这一假说,我们使用冷冻电子断层扫描来检查来自 BVM 处理细胞的(非感染性)HIV-1 病毒粒子的结构。我们发现这些粒子包含一个不完整的密度壳,位于病毒包膜下,具有类似于不成熟 HIV 的 Gag 晶格的六边形蜂窝结构,但缺乏最内层、与 NC 相关的层。我们得出结论,该壳代表已加工的不成熟 Gag 晶格的残留物,但基本上保持完整,除了 CA-SP1 位点。我们还将 BVM 处理的粒子与突变 CA5 形成的病毒粒子进行了比较,在 CA5 中,CA 和 SP1 之间的切割也被阻断。在这里,我们发现 CA 相关的壳更薄,没有可见的蜂窝组织的迹象,表明构象发生了改变,进一步表明 BVM 的结合稳定了不成熟的晶格。在这两种情况下,观察到的未能组装成熟衣壳与丧失感染力相关。