Impact of subtype C‑specific amino acid variants on HIV‑1 Tat‑TAR interaction: insights from molecular modelling and dynamics

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