Non-active site mutants of HIV-1 protease influence resistance and sensitisation towards protease inhibitors

Abstract

Background: HIV-1 can develop resistance to antiretroviral drugs, mainly through mutations within the target regions of the drugs. In HIV-1 protease, a majority of resistance-associated mutations that develop in response to therapy with protease inhibitors are found in the protease's active site that serves also as a binding pocket for the protease inhibitors, thus directly impacting the protease-inhibitor interactions. Some resistance-associated mutations, however, are found in more distant regions, and the exact mechanisms how these mutations affect protease-inhibitor interactions are unclear. Furthermore, some of these mutations, e.g. N88S and L76V, do not only induce resistance to the currently administered drugs, but contrarily induce sensitivity towards other drugs. In this study, mutations N88S and L76V, along with three other resistance-associated mutations, M46I, I50L, and I84V, are analysed by means of molecular dynamics simulations to investigate their role in complexes of the protease with different inhibitors and in different background sequence contexts. Results: Using these simulations for alchemical calculations to estimate the effects of mutations M46I, I50L, I84V, N88S, and L76V on binding free energies shows they are in general in line with the mutations' effect on IC 50 values. For the primary mutation L76V, however, the presence of a background mutation M46I in our analysis influences whether the unfavourable effect of L76V on inhibitor binding is sufficient to outweigh the accompanying reduction in catalytic activity of the protease. Finally, we show that L76V and N88S changes the hydrogen bond stability of these residues with residues D30/K45 and D30/T31/T74, respectively. Conclusions: We demonstrate that estimating the effect of both binding pocket and distant mutations on inhibitor binding free energy using alchemical calculations can reproduce their effect on the experimentally measured IC 50 values. We show that distant site mutations L76V and N88S affect the hydrogen bond network in the protease's active site, which offers an explanation for the indirect effect of these mutations on inhibitor binding. This work thus provides valuable insights on interplay between primary and background mutations and mechanisms how they affect inhibitor binding.

One Plus One Makes Three: Triangular Coupling of Correlated Amino Acid Mutations

“…However, several studies showed that some thermodynamic couplings persist over large distances in space, which can not be explained by direct interactions of the corresponding amino acids. − The long-range mutation effects in proteins are of particular interest as they may have significant contributions to enzymatic activity, , protein folding, or ligand binding affinity . In coevolution analyses coupled mutations can be traced down by multiple sequence alignments.…”

Acquired HIV-1 Protease Conformational Flexibility Associated with Lopinavir Failure May Shape the Outcome of Darunavir Therapy after Antiretroviral Therapy Switch

“…Earlier studies have shown that intrinsic changes in the HIV-1 PR conformation associated with the accumulation of drug-resistant mutations may cause geometric alteration in the HIV-1 PR structure affecting the active site and other domains [61,77]. These intrinsic changes may cause sensitization and cross-resistance to HIV-1 PIs by altering the molecular interaction of the former with the protein during binding [16,77]. The evolution of these characteristics in proteins is often associated with the loss of protein stability [80].…”

“…The gain in HIV-1 PR flexibility due to changes in inter-residue Drug-resistance mutations could be found in the active site of HIV-1 PR and directly impact the binding affinity and interaction of HIV-1 PIs with HIV-1 PR [15]. In contrast, non-active site mutations may not directly affect the interaction of HIV-1 PR with inhibitors but may indirectly influence the molecular interaction of inhibitors with the HIV-1 PR through alteration of the protein flexibility and stability [16][17][18]. The accumulation and interplay between active and non-active site drug-resistance mutations arising from drug pressure may cause structural changes, leading to HIV-1 PR variants with altered protein conformations [19,20].…”

Identification of Aloe-derived natural products as prospective lead scaffolds for SARS-CoV-2 main protease (Mpro) inhibitors

“…1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 17 , 18 , 19 , 20 , 21 , 22 , 31 , 32 , 33 , 34 , 35 , 36 , 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 , 51 , 52 , 53 , 54 , 55 , 56 , 57 , 58 , 59 , 60 , 61 , 63 , 64 .…”