pRetroviral proteases have been shown previously to be only active as homodimers. They are essential to form the separate and active proteins from the viral precursors. Spumaretroviruses produce separate precursors for Gag and Pol, rather than a Gag and a Gag–Pol precursor. Nevertheless, processing of Pol into a PR (protease)–RT (reverse transcriptase) and integrase is essential in order to obtain infectious viral particles. We showed recently that the PR–RT from a simian foamy virus, as well as the separate PRshort (protease) domain, exhibit proteolytic activities, although only monomeric forms could be detected. In the present study, we demonstrate that PRshort and PR–RT can be inhibited by the putative dimerization inhibitor cholic acid. Various other inhibitors, including darunavir and tipranavir, known to prevent HIV-1 PR dimerization in cells, had no effect on foamy virus protease iin vitro/i. sup1/supH-sup15/supN HSQC (heteronuclear single quantum coherence) NMR analysis of PRshort indicates that cholic acid binds in the proposed PRshort dimerization interface and appears to impair formation of the correct dimer. NMR analysis by paramagnetic relaxation enhancement resulted in elevated transverse relaxation rates of those amino acids predicted to participate in dimer formation. Our results suggest transient PRshort homodimers are formed under native conditions but are only present as a minor transient species, which is not detectable by traditional methods./p
展开▼
机译：先前已经显示逆转录病毒蛋白酶仅作为同型二聚体具有活性。它们对于与病毒前体形成分离的活性蛋白至关重要。斑驳病毒产生Gag和Pol的单独前体，而不是Gag和Gag-Pol的前体。然而，为了获得感染性病毒颗粒，将Pol加工成PR（蛋白酶）-RT（逆转录酶）和整合酶是必不可少的。我们最近表明，来自猿猴泡沫病毒的PR–RT以及单独的PRshort（蛋白酶）结构域均具有蛋白水解活性，尽管只能检测到单体形式。在本研究中，我们证明了PRshort和PR–RT可被假定的二聚化抑制剂胆酸抑制。已知能阻止细胞中HIV-1 PR二聚化的其他各种抑制剂，包括darunavir和Tipranavir，在体外对泡沫病毒蛋白酶没有影响。 PRshort的 1 H- 15 N HSQC（异核单量子相干）NMR分析表明，胆酸结合在拟议的PRshort二聚化界面中，似乎损害了正确二聚体的形成。通过顺磁弛豫增强进行的NMR分析导致预测参与二聚体形成的那些氨基酸的横向弛豫速率升高。我们的结果表明，短暂的PRshort同型二聚体是在自然条件下形成的，但仅作为次要的短暂物种存在，这是传统方法无法检测到的。
展开▼
