Sun Qinfang, Biswas Avik, Lyumkis Dmitry, Levy Ronald, Deng Nanjie
Center for Biophysics and Computational Biology and Department of Chemistry, Temple University, Philadelphia, PA 19122.
The Salk Institute for Biological Studies, Laboratory of Genetics, La Jolla, CA 92037.
bioRxiv. 2023 Dec 1:2023.11.29.569269. doi: 10.1101/2023.11.29.569269.
The first and second-generation clinically used HIV-1 integrase (IN) strand transfer inhibitors (INSTIs) are key components of antiretroviral therapy (ART), which work by blocking the integration step in the HIV-1 replication cycle that is catalyzed by a nucleoprotein assembly called an intasome. However, resistance to even the latest clinically used INSTIs is beginning to emerge. Developmental third-generation INSTIs, based on naphthyridine scaffold, are promising candidates to combat drug-resistant viral variants. Among these novel INSTIs, compound 4f exhibits two distinct conformations when binding to intasomes from HIV-1 and the closely related prototype foamy virus (PFV), despite the high structural similarity of their INSTI binding pockets. The molecular mechanism and the key active site residues responsible for these differing binding modes in closely related intasomes remain elusive. To unravel the molecular determinants governing the two distinct binding modes, we employ a novel molecular dynamics-based free energy approach that utilizes alchemical pathways to overcome the sampling challenges associated with transitioning between two ligand conformations within crowded environments along physical pathways. The calculated conformational free energies successfully recapitulate the experimentally observed binding mode preferences in the two viral intasomes. Analysis of the simulated structures suggests that the observed binding mode preferences are caused by amino acid residue differences in both the front and the central catalytic sub-pocket of the INSTI binding site in HIV-1 and PFV. Additional free energy calculations on mutants of HIV-1 and PFV revealed that while both sub-pockets contribute to the binding mode selection, the central sub-pocket plays a more important role. These results highlight the importance of both side chain and solvent reorganization, as well as the conformational entropy in determining the ligand binding mode and will help inform the development of more effective INSTIs for combatting drug-resistant viral variants.
第一代和第二代临床上使用的HIV-1整合酶（IN）链转移抑制剂（INSTIs）是抗逆转录病毒疗法（ART）的关键组成部分，其作用机制是阻断HIV-1复制周期中的整合步骤，该步骤由一种称为整合体的核蛋白组装体催化。然而，即使是最新的临床使用的INSTIs也开始出现耐药性。基于萘啶支架的第三代INSTIs正在开发中，有望成为对抗耐药病毒变体的候选药物。在这些新型INSTIs中，化合物4f在与HIV-1和密切相关的原型泡沫病毒（PFV）的整合体结合时表现出两种不同的构象，尽管它们的INSTI结合口袋在结构上高度相似。在密切相关的整合体中，导致这些不同结合模式的分子机制和关键活性位点残基仍然不清楚。为了揭示控制这两种不同结合模式的分子决定因素，我们采用了一种基于分子动力学的新型自由能方法，该方法利用炼金术途径来克服与沿着物理途径在拥挤环境中两种配体构象之间转换相关的采样挑战。计算得到的构象自由能成功地重现了在两种病毒整合体中实验观察到的结合模式偏好。对模拟结构的分析表明，观察到的结合模式偏好是由HIV-1和PFV中INSTI结合位点的前部和中央催化亚口袋中的氨基酸残基差异引起的。对HIV-1和PFV突变体的额外自由能计算表明，虽然两个亚口袋都有助于结合模式的选择，但中央亚口袋起着更重要的作用。这些结果突出了侧链和溶剂重组以及构象熵在确定配体结合模式中的重要性，并将有助于为开发更有效的INSTIs以对抗耐药病毒变体提供信息。
