Depollier Julien, Hourdou Marie-Laure, Aldrian-Herrada Gudrun, Rothwell Paul, Restle Tobias, Divita Gilles
Department of Molecular Biophysics & Therapeutics, Centre de Recherches de Biochimie Macromoléculaire, FRE-2593 CNRS, 1919 Route de Mende, 34293 Montpellier, France.
Biochemistry. 2005 Feb 15;44(6):1909-18. doi: 10.1021/bi0484264.
The biologically active forms of human immunodeficiency viruses type 1 and 2 reverse transcriptase (RT) found in infectious virions are heterodimers. We have previously shown that the dimeric nature of reverse transcriptase represents an important target for the design of a new class of antiviral agents and have designed a short peptide (Pep-7) derived from the tryptophan-rich motif of the connection subdomain that blocks dimerization of reverse transcriptase in vitro and abolishes viral infection. In the present work, we have investigated the mechanism through which this peptide inhibits RT dimerization and consequently viral propagation. We demonstrate that Pep-7 interacts preferentially with the p51 subunit within the heterodimeric reverse transcriptase, which destabilizes reverse transcriptase dimer conformation, thereby triggering dissociation. We have identified two residues Trp(24) and Phe(61), located on the fingers subdomain of p51, required for Pep-7 binding. Selective mutation of these residues on p51 to a glycine dramatically alters the stability of the RT-heterodimer suggesting that the fingers subdomain of p51 is also involved in stabilization of reverse transcriptase. We propose that the binding site of Pep-7 is located in a cleft between the fingers and the connection subdomains of p51 that contains the two highly conserved residues Phe(61) and Trp(24).
在感染性病毒粒子中发现的1型和2型人类免疫缺陷病毒逆转录酶（RT）的生物活性形式是异二聚体。我们之前已经表明，逆转录酶的二聚体性质是设计新型抗病毒药物的一个重要靶点，并且我们设计了一种源自连接亚结构域富含色氨酸基序的短肽（Pep-7），它在体外可阻止逆转录酶的二聚化并消除病毒感染。在本研究中，我们研究了该肽抑制RT二聚化并进而抑制病毒传播的机制。我们证明Pep-7优先与异二聚体逆转录酶中的p51亚基相互作用，这会破坏逆转录酶二聚体的构象，从而引发解离。我们确定了位于p51的指状亚结构域上的两个残基Trp(24)和Phe(61)，它们是Pep-7结合所必需的。将p51上的这些残基选择性突变为甘氨酸会显著改变RT异二聚体的稳定性，这表明p51的指状亚结构域也参与逆转录酶的稳定。我们提出Pep-7的结合位点位于p51的指状亚结构域和连接亚结构域之间的一个裂隙中，该裂隙包含两个高度保守的残基Phe(61)和Trp(24)。
