Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15260, USA.
Chem Biol Drug Des. 2012 Nov;80(5):706-16. doi: 10.1111/cbdd.12010. Epub 2012 Aug 31.
HIV-1 reverse transcriptase (RT) has been an attractive target for the development of antiretroviral agents. Although this enzyme is bi-functional, having both DNA polymerase and ribonuclease H (RNH) activities, there is no clinically approved inhibitor of the RNH activity. Here, we characterize the structural basis and molecular interaction of an allosteric site inhibitor, BHMP07, with the wild-type (WT) RNH fragment. Solution NMR experiments for inhibitor titration on WT RNH showed relatively wide chemical shift perturbations, suggesting a long-range conformational effect on the inhibitor interaction. Comparisons of the inhibitor-induced NMR chemical shift changes of RNH with those of RNH dimer, in the presence and absence of Mg(2+) , were performed to determine and verify the interaction site. The NMR results, with assistance of molecular docking, indicate that BHMP07 preferentially binds to a site that is located between the RNH active site and the region encompassing helices B and D (the 'substrate-handle region'). The interaction site is consistent with the previous proposed site, identified using a chimeric RNH (p15-EC) [Gong et al. (2011) Chem Biol Drug Des 77, 39-47], but with slight differences that reflect the characteristics of the amino acid sequences in p15-EC compared to the WT RNH.
HIV-1 逆转录酶（RT）一直是开发抗逆转录病毒药物的有吸引力的靶标。尽管该酶具有双功能，具有 DNA 聚合酶和核糖核酸酶 H（RNH）活性，但尚未有临床批准的 RNH 活性抑制剂。在这里，我们描述了变构位点抑制剂 BHMP07 与野生型（WT）RNH 片段的结构基础和分子相互作用。用于 WT RNH 抑制剂滴定的溶液 NMR 实验显示相对较宽的化学位移扰动，表明对抑制剂相互作用具有远程构象效应。在存在和不存在 Mg2+的情况下，比较了抑制剂诱导的 RNH 的 NMR 化学位移变化与 RNH 二聚体的变化，以确定和验证相互作用位点。NMR 结果，在分子对接的帮助下，表明 BHMP07 优先结合位于 RNH 活性位点和包含螺旋 B 和 D 的区域（“底物手柄区域”）之间的位点。该相互作用位点与先前使用嵌合 RNH（p15-EC）[Gong 等人（2011）Chem Biol Drug Des 77, 39-47]确定的提议位点一致，但存在一些差异，这些差异反映了 p15-EC 与 WT RNH 相比氨基酸序列的特征。
