Small molecules targeted to a non-catalytic "RVxF" binding site of protein phosphatase-1 inhibit HIV-1.
Small molecules targeted to a non-catalytic "RVxF" binding site of protein phosphatase-1 inhibit HIV-1.
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DOI：
10.1371/journal.pone.0039481
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Nekhai S
中科院分区：
文献类型：
--
作者：
Ammosova T;Platonov M;Yedavalli VR;Obukhov Y;Gordeuk VR;Jeang KT;Kovalskyy D;Nekhai S
HIV-1达特蛋白募集宿主细胞因子，包括CDK 9/细胞周期蛋白T1至HIV-1 TAR RNA，从而诱导HIV-1转录。与宿主Ser/Thr蛋白磷酸酶-1（PP 1）的相互作用对于达特的这一功能至关重要。PP 1与达特序列Q35 VCF 38结合，Q35 VCF 38类似于存在于PP 1结合调节亚基上的PP 1结合“RNF”基序。我们发现，PP 1结合肽的表达，一个核心结构域的核抑制剂的PP 1，破坏HIV-1的达特与PP 1的相互作用，抑制HIV-1的转录和复制。在这里，我们报告的小分子化合物，目标的“RIF”结合腔的PP 1破坏的相互作用的PP 1与达特和抑制HIV-1复制。利用PP 1的晶体结构，我们实际上筛选了30万种化合物，并鉴定了262种预计与PP 1的“RIF”容纳腔结合的小分子。然后测定这些化合物对CEM T细胞中HIV-1转录的抑制。其中一种化合物1H 4在非细胞毒性浓度下抑制HIV-1的转录和复制。1H 4在体外阻止PP 1介导的含有RNF序列的底物肽的去磷酸化。1H 4还破坏了培养细胞中PP 1与达特的结合，而对PP 1与细胞调节因子NIPP 1和PNUTS的相互作用或对细胞蛋白质组没有影响。最后，1H 4阻止PP 1易位到细胞核。总之，我们的研究表明，HIV抑制可以通过使用小分子靶向PP 1的非催化位点来实现。这项原理验证研究可以作为开发新型抗病毒药物的起点，这些药物靶向HIV-1病毒蛋白与其宿主伴侣的界面。
HIV-1 Tat protein recruits host cell factors including CDK9/cyclin T1 to HIV-1 TAR RNA and thereby induces HIV-1 transcription. An interaction with host Ser/Thr protein phosphatase-1 (PP1) is critical for this function of Tat. PP1 binds to a Tat sequence, Q35VCF38, which resembles the PP1-binding “RVxF” motif present on PP1-binding regulatory subunits. We showed that expression of PP1 binding peptide, a central domain of Nuclear Inhibitor of PP1, disrupted the interaction of HIV-1 Tat with PP1 and inhibited HIV-1 transcription and replication. Here, we report small molecule compounds that target the “RVxF”-binding cavity of PP1 to disrupt the interaction of PP1 with Tat and inhibit HIV-1 replication. Using the crystal structure of PP1, we virtually screened 300,000 compounds and identified 262 small molecules that were predicted to bind the “RVxF”-accommodating cavity of PP1. These compounds were then assayed for inhibition of HIV-1 transcription in CEM T cells. One of the compounds, 1H4, inhibited HIV-1 transcription and replication at non-cytotoxic concentrations. 1H4 prevented PP1-mediated dephosphorylation of a substrate peptide containing an RVxF sequence in vitro. 1H4 also disrupted the association of PP1 with Tat in cultured cells without having an effect on the interaction of PP1 with the cellular regulators, NIPP1 and PNUTS, or on the cellular proteome. Finally, 1H4 prevented the translocation of PP1 to the nucleus. Taken together, our study shows that HIV- inhibition can be achieved through using small molecules to target a non-catalytic site of PP1. This proof-of-principle study can serve as a starting point for the development of novel antiviral drugs that target the interface of HIV-1 viral proteins with their host partners.
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