Probing the "fingers" domain binding pocket of Hepatitis C virus NS5B RdRp and D559G resistance mutation via molecular docking, molecular dynamics simulation and binding free energy calculations
Probing the "fingers" domain binding pocket of Hepatitis C virus NS5B RdRp and D559G resistance mutation via molecular docking, molecular dynamics simulation and binding free energy calculations
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DOI：
10.1080/07391102.2018.1491419
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发表时间：
2019-06-13
影响因子：
4.4
通讯作者：
Kumaradhas, Poomani
中科院分区：
文献类型：
--
作者：
Manjula, Saravanan;Sivanandam, Magudeeswaran;Kumaradhas, Poomani
NS5B RdRp聚合酶是丙型肝炎病毒复制的重要酶。在丙型肝炎病毒复制过程中，模板RNA结合发生在NS5B的“Finger”亚域中。“Finger”结构域是丙型肝炎病毒药物开发的一个新的变构位点。“Finger”亚区的抑制剂采用了一种新的抗病毒机制，称为RNA干预。从必需氨基酸残基、配体的结合方式以及RNA干预的活性部位分子间相互作用的细节可以看出，这一机制在实验研究中是模糊的。为了阐明这些细节，我们进行了Finger结构域抑制剂quercetagetin(QGN)与NS5B聚合酶的分子对接分析。对QGN-NS5B的分子间相互作用进行了详细的分析，发现QGN与结合口袋氨基酸残基Ala97、Ala140、Ile160、Phe162、Gly283、Gly557和Asp559相互作用，并与Phe162形成pi中线水平椭圆pi堆积作用，与Gly283形成氢键作用。这些都被发现是RNA干预机制的基本相互作用。在较强的氢键相互作用中，QGNMIDLINE水平ELLIPSISAla140是本工作新发现的一种重要的氢键相互作用，这种相互作用没有被以前报道的晶体结构所解析。由于D559G突变导致QGN的抑制率降低到7倍，因此我们对野生和D559G突变的复合体进行了分子动力学模拟，以研究蛋白质构象的稳定性和分子间的相互作用。在50 ns MD模拟结束时，Phe162与QGN在野生型复合体中发现的pi中线水平椭圆pi堆积作用转变为T形pi堆积作用，从而降低了抑制强度。采用分子动力学模拟、结合自由能计算和主成分分析相结合的方法对D559G抗性突变的起源进行了研究。将结果与野生型复合体进行了比较。突变D559G降低了QGN分子与Finger结构域的结合亲和力。对野生型和突变复合体的每个残基的自由能分解分析表明，非极性能量贡献的损失是抗性的根源。
The NS5B RdRp polymerase is a prominent enzyme for the replication of Hepatitis C virus (HCV). During the HCV replication, the template RNA binding takes place in the "fingers" sub-domain of NS5B. The "fingers" domain is a new emerging allosteric site for the HCV drug development. The inhibitors of the "fingers" sub-domain adopt a new antiviral mechanism called RNA intervention. The details of essential amino acid residues, binding mode of the ligand, and the active site intermolecular interactions of RNA intervention reflect that this mechanism is ambiguous in the experimental study. To elucidate these details, we performed molecular docking analysis of the fingers domain inhibitor quercetagetin (QGN) with NS5B polymerase. The detailed analysis of QGN-NS5B intermolecular interactions was carried out and found that QGN interacts with the binding pocket amino acid residues Ala97, Ala140, Ile160, Phe162, Gly283, Gly557, and Asp559; and also forms pi MIDLINE HORIZONTAL ELLIPSIS pi stacking interaction with Phe162 and hydrogen bonding interaction with Gly283. These are found to be the essential interactions for the RNA intervention mechanism. Among the strong hydrogen bonding interactions, the QGNMIDLINE HORIZONTAL ELLIPSISAla140 is a newly identified important hydrogen bonding interaction by the present work and this interaction was not resolved by the previously reported crystal structure. Since D559G mutation at the fingers domain was reported for reducing the inhibition percentage of QGN to sevenfold, we carried out molecular dynamics (MD) simulation for wild and D559G mutated complexes to study the stability of protein conformation and intermolecular interactions. At the end of 50 ns MD simulation, the pi MIDLINE HORIZONTAL ELLIPSIS pi stacking interaction of Phe162 with QGN found in the wild-type complex is altered into T-shaped pi stacking interaction, which reduces the inhibition strength. The origin of the D559G resistance mutation was studied using combined MD simulation, binding free energy calculations and principal component analysis. The results were compared with the wild-type complex. The mutation D559G reduces the binding affinity of the QGN molecule to the fingers domain. The free energy decomposition analysis of each residue of wild-type and mutated complexes revealed that the loss of non-polar energy contribution is the origin of the resistance.