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Abstract：
BACKGROUND: HIV-1 produces Tat, a crucial protein for transcription, viral replication, and CNS neurotoxicity. Tat interacts with TAR, enhancing HIV reverse transcription. Subtype C Tat variants (C31S, R57S, Q63E) are associated with reduced transactivation and neurovirulence compared to subtype B. However, their precise impact on Tat-TAR binding is unclear. This study investigates how these substitutions affect Tat-TAR interaction.
METHODS: We utilized molecular modelling techniques, including MODELLER, to produce precise three-dimensional structures of HIV-1 Tat protein variants. We utilized Tat subtype B as the reference or wild type, and generated Tat variants to mirror those amino acid variants found in Tat subtype C. Subtype C-specific amino acid substitutions were selected based on their role in the neuropathogenesis of HIV-1. Subsequently, we conducted molecular docking of each Tat protein variant to TAR using HDOCK, followed by molecular dynamic simulations.
RESULTS: Molecular docking results indicated that Tat subtype B (TatWt) showed the highest affinity for the TAR element (-262.07), followed by TatC31S (-261.61), TatQ63E (-256.43), TatC31S/R57S/Q63E (-238.92), and TatR57S (-222.24). However, binding free energy analysis showed higher affinities for single variants TatQ63E (-349.2 ± 10.4 kcal/mol) and TatR57S (-290.0 ± 9.6 kcal/mol) compared to TatWt (-247.9 ± 27.7 kcal/mol), while TatC31S and TatC31S/R57SQ/63E showed lower values. Interactions over the protein trajectory were also higher for TatQ63E and TatR57S compared to TatWt, TatC31S, and TatC31S/R57SQ/63E, suggesting that modifying amino acids within the Arginine/Glutamine-rich region notably affects TAR interaction. Single amino acid mutations TatR57S and TatQ63E had a significant impact, while TatC31S had minimal effect. Introducing single amino acid variants from TatWt to a more representative Tat subtype C (TatC31S/R57SQ/63E) resulted in lower predicted binding affinity, consistent with previous findings.
CONCLUSIONS: These identified amino acid positions likely contribute significantly to Tat-TAR interaction and the differential pathogenesis and neuropathogenesis observed between subtype B and subtype C. Additional experimental investigations should prioritize exploring the influence of these amino acid signatures on TAR binding to gain a comprehensive understanding of their impact on viral transactivation, potentially identifying them as therapeutic targets.
METHODS: We utilized molecular modelling techniques, including MODELLER, to produce precise three-dimensional structures of HIV-1 Tat protein variants. We utilized Tat subtype B as the reference or wild type, and generated Tat variants to mirror those amino acid variants found in Tat subtype C. Subtype C-specific amino acid substitutions were selected based on their role in the neuropathogenesis of HIV-1. Subsequently, we conducted molecular docking of each Tat protein variant to TAR using HDOCK, followed by molecular dynamic simulations.
RESULTS: Molecular docking results indicated that Tat subtype B (TatWt) showed the highest affinity for the TAR element (-262.07), followed by TatC31S (-261.61), TatQ63E (-256.43), TatC31S/R57S/Q63E (-238.92), and TatR57S (-222.24). However, binding free energy analysis showed higher affinities for single variants TatQ63E (-349.2 ± 10.4 kcal/mol) and TatR57S (-290.0 ± 9.6 kcal/mol) compared to TatWt (-247.9 ± 27.7 kcal/mol), while TatC31S and TatC31S/R57SQ/63E showed lower values. Interactions over the protein trajectory were also higher for TatQ63E and TatR57S compared to TatWt, TatC31S, and TatC31S/R57SQ/63E, suggesting that modifying amino acids within the Arginine/Glutamine-rich region notably affects TAR interaction. Single amino acid mutations TatR57S and TatQ63E had a significant impact, while TatC31S had minimal effect. Introducing single amino acid variants from TatWt to a more representative Tat subtype C (TatC31S/R57SQ/63E) resulted in lower predicted binding affinity, consistent with previous findings.
CONCLUSIONS: These identified amino acid positions likely contribute significantly to Tat-TAR interaction and the differential pathogenesis and neuropathogenesis observed between subtype B and subtype C. Additional experimental investigations should prioritize exploring the influence of these amino acid signatures on TAR binding to gain a comprehensive understanding of their impact on viral transactivation, potentially identifying them as therapeutic targets.
摘要：
背景：HIV-1产生Tat，转录的关键蛋白质，病毒复制，和中枢神经系统神经毒性。Tat与TAR互动，增强HIV逆转录。亚型CTat变体（C31S，R57S,与B亚型相比，Q63E）与降低的反式激活和神经毒力有关。它们对Tat-TAR结合的确切影响尚不清楚。本研究调查了这些替代如何影响Tat-TAR相互作用。
方法：我们利用分子建模技术，包括Modeller,以产生HIV-1Tat蛋白变体的精确三维结构。我们使用Tat亚型B作为参考或野生型，并产生Tat变体以反映在Tat亚型C中发现的那些氨基酸变体。亚型C特异性氨基酸取代是基于它们在HIV-1的神经发病机理中的作用而选择的。随后，我们使用HDOCK将每个Tat蛋白变体与TAR进行分子对接，其次是分子动力学模拟。
结果：分子对接结果表明，Tat亚型B（TatWt）对TAR元件的亲和力最高（-262.07），其次是TatC31S（-261.61），TatQ63E(-256.43),TatC31S/R57S/Q63E(-238.92),和TatR57S（-222.24）。然而,结合自由能分析显示，与TatWt（-247.9±27.7kcal/mol）相比，单个变体TatQ63E（-349.2±10.4kcal/mol）和TatR57S（-290.0±9.6kcal/mol）的亲和力更高，而TatC31S和TatC31S/R57SQ/63E显示较低的值。与TatWt相比，TatQ63E和TatR57S在蛋白质轨迹上的相互作用也更高。TatC31S,和TatC31S/R57SQ/63E，这表明修饰富含精氨酸/谷氨酰胺的区域内的氨基酸显着影响TAR相互作用。单氨基酸突变TatR57S和TatQ63E有显著影响,而TatC31S影响最小。将单个氨基酸变体从TatWt引入更具代表性的Tat亚型C（TatC31S/R57SQ/63E）导致较低的预测结合亲和力，与以前的发现一致。
结论：这些确定的氨基酸位置可能对Tat-TAR相互作用以及B亚型和C亚型之间观察到的差异发病机制和神经发病机制有重要贡献。其他实验研究应优先探索这些氨基酸特征对TAR结合的影响，以全面了解它们对病毒反式激活的影响。可能将它们确定为治疗靶标。
方法：我们利用分子建模技术，包括Modeller,以产生HIV-1Tat蛋白变体的精确三维结构。我们使用Tat亚型B作为参考或野生型，并产生Tat变体以反映在Tat亚型C中发现的那些氨基酸变体。亚型C特异性氨基酸取代是基于它们在HIV-1的神经发病机理中的作用而选择的。随后，我们使用HDOCK将每个Tat蛋白变体与TAR进行分子对接，其次是分子动力学模拟。
结果：分子对接结果表明，Tat亚型B（TatWt）对TAR元件的亲和力最高（-262.07），其次是TatC31S（-261.61），TatQ63E(-256.43),TatC31S/R57S/Q63E(-238.92),和TatR57S（-222.24）。然而,结合自由能分析显示，与TatWt（-247.9±27.7kcal/mol）相比，单个变体TatQ63E（-349.2±10.4kcal/mol）和TatR57S（-290.0±9.6kcal/mol）的亲和力更高，而TatC31S和TatC31S/R57SQ/63E显示较低的值。与TatWt相比，TatQ63E和TatR57S在蛋白质轨迹上的相互作用也更高。TatC31S,和TatC31S/R57SQ/63E，这表明修饰富含精氨酸/谷氨酰胺的区域内的氨基酸显着影响TAR相互作用。单氨基酸突变TatR57S和TatQ63E有显著影响,而TatC31S影响最小。将单个氨基酸变体从TatWt引入更具代表性的Tat亚型C（TatC31S/R57SQ/63E）导致较低的预测结合亲和力，与以前的发现一致。
结论：这些确定的氨基酸位置可能对Tat-TAR相互作用以及B亚型和C亚型之间观察到的差异发病机制和神经发病机制有重要贡献。其他实验研究应优先探索这些氨基酸特征对TAR结合的影响，以全面了解它们对病毒反式激活的影响。可能将它们确定为治疗靶标。
