Conformational state of a 25-mer peptide from the cyclophilin-binding loop of the HIV type 1 capsid protein.
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ABSTRACT:
SUBMITTER: Reimer U
PROVIDER: S-EPMC1218652 | biostudies-other | 1997 Aug
REPOSITORIES: biostudies-other
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Project description:The human myxovirus-resistance protein B (MxB, also called Mx2) was recently reported to inhibit HIV-1 infection by impeding the nuclear import and integration of viral DNA. However, it is currently unknown whether there exist MxB-resistant HIV-1 strains in the infected individuals. Answer to this question should address whether MxB exerts an inhibitory pressure on HIV-1 in vivo and whether HIV-1 has evolved to evade MxB inhibition.We have examined ten transmitted founder (T/F) HIV-1 strains for their sensitivity to MxB inhibition by infecting CD4+ T cell lines SupT1 and PM1 that were stably transduced to express MxB. Two T/F stains, CH040.c and RHPA.c, were found resistant and this resistance phenotype was mapped to the amino acid positions 87 and 208 in viral capsid. The H87Q mutation is located in the cyclophilin A (CypA) binding loop and has a prevalence of 21% in HIV-1 sequences registered in HIV database. This finding prompted us to test other frequent amino acid variants in the CypA-binding region and the results revealed MxB-resistant mutations at amino acid positions 86, 87, 88 and 92 in capsid. All these mutations diminished the interaction of HIV-1 capsid with CypA.Our results demonstrate the existence of MxB-resistant T/F HIV-1 strains. The high prevalence of MxB-resistant mutations in the CypA-binding loop indicates the significant selective pressure of MxB on HIV-1 replication in vivo especially given that this viral resistance mechanism operates at expense of losing CypA.
| S-EPMC4300040 | biostudies-literature
Project description:Host factor protein Cyclophilin A (CypA) regulates HIV-1 viral infectivity through direct interactions with the viral capsid, by an unknown mechanism. CypA can either promote or inhibit viral infection, depending on host cell type and HIV-1 capsid (CA) protein sequence. We have examined the role of conformational dynamics on the nanosecond to millisecond timescale in HIV-1 CA assemblies in the escape from CypA dependence, by magic-angle spinning (MAS) NMR and molecular dynamics (MD). Through the analysis of backbone (1)H-(15)N and (1)H-(13)C dipolar tensors and peak intensities from 3D MAS NMR spectra of wild-type and the A92E and G94D CypA escape mutants, we demonstrate that assembled CA is dynamic, particularly in loop regions. The CypA loop in assembled wild-type CA from two strains exhibits unprecedented mobility on the nanosecond to microsecond timescales, and the experimental NMR dipolar order parameters are in quantitative agreement with those calculated from MD trajectories. Remarkably, the CypA loop dynamics of wild-type CA HXB2 assembly is significantly attenuated upon CypA binding, and the dynamics profiles of the A92E and G94D CypA escape mutants closely resemble that of wild-type CA assembly in complex with CypA. These results suggest that CypA loop dynamics is a determining factor in HIV-1's escape from CypA dependence.
| S-EPMC4664340 | biostudies-literature
Project description:The cellular protein, cyclophilin A (CypA), is incorporated into the virion of the type 1 human immunodeficiency virus (HIV-1) via a direct interaction with the capsid domain of the viral Gag polyprotein. We demonstrate that the capsid sequence 87His-Ala-Gly-Pro-Ile-Ala92 (87HAGPIA92) encompasses the primary cyclophilin A binding site and present an X-ray crystal structure of the CypA/HAGPIA complex. In contrast to the cis prolines observed in all previously reported structures of CypA complexed with model peptides, the proline in this peptide, Pro 90, binds the cyclophilin A active site in a trans conformation. We also report the crystal structure of a complex between CypA and the hexapeptide HVGPIA, which also maintains the trans conformation. Comparison with the recently determined structures of CypA in complexes with larger fragments of the HIV-1 capsid protein demonstrates that CypA recognition of these hexapeptides involves contacts with peptide residues Ala(Val) 88, Gly 89, and Pro 90, and is independent of the context of longer sequences.
| S-EPMC2143596 | biostudies-other
Project description:SARS-CoV-2 is the novel coronavirus causing the COVID-19 pandemic. To enter human cells, the receptor-binding domain (RBD) of the S1 subunit of SARS-CoV-2 (SARS-CoV-2-RBD) initially binds to the peptidase domain of angiotensin-converting enzyme 2 receptor (ACE2-PD). Using peptides to inhibit SARS-CoV-2-RBD binding to ACE2 is a potential therapeutic solution for COVID-19. A previous study identified a 23-mer peptide (SBP1) that bound to SARS-CoV-2-RBD with comparable KD to ACE2. We employed computational protein design and molecular dynamics (MD) to design SARS-CoV-2-RBD 25-mer peptide binders (SPB25) with better predicted binding affinity than SBP1. Using residues 21-45 of the α1 helix of ACE2-PD as the template, our strategy is employing Rosetta to enhance SPB25 binding affinity to SARS-CoV-2-RBD and avoid disrupting existing favorable interactions by using residues that have not been reported to form favorable interactions with SARS-CoV-2-RBD as designed positions. Designed peptides with better predicted binding affinities, by Rosetta, than SPB25 were subjected to MD validation. The MD results show that five designed peptides (SPB25F8N, SPB25F8R, SPB25L25R, SPB25F8N/L25R, and SPB25F8R/L25R) have better predicted binding affinities, by the MM-GBSA method, than SPB25 and SBP1. This study developed an approach to design SARS-CoV-2-RBD peptide binders, and these peptides may be promising candidates as potential SARS-CoV-2 inhibitors.
| S-EPMC7687296 | biostudies-literature
Project description:Understanding structural transitions within macromolecules remains an important challenge in biochemistry, with important implications for drug development and medicine. Insight into molecular behavior often requires residue-specific dynamics measurement at micromolar concentrations. We studied MP01-Gen4, a library peptide selected to rapidly undergo bioconjugation, by using electron paramagnetic resonance (EPR) to measure conformational dynamics. We mapped the dynamics of MP01-Gen4 with residue-specificity and identified the regions involved in a structural transformation related to the conjugation reaction. Upon reaction, the conformational dynamics of residues near the termini slow significantly more than central residues, indicating that the reaction induces a structural transition far from the reaction site. Arrhenius analysis demonstrates a nearly threefold decrease in the activation energy of conformational diffusion upon reaction (8.0 kBT to 3.4 kBT), which occurs across the entire peptide, independently of residue position. This novel approach to EPR spectral analysis provides insight into the positional extent of disorder and the nature of the energy landscape of a highly reactive, intrinsically disordered library peptide before and after conjugation.
| S-EPMC7018720 | biostudies-literature
Project description:We have studied the effects associated with two single amino acid substitution mutations in HIV-1 capsid (CA), the E98A and E187G. Both amino acids are well conserved among all major HIV-1 subtypes. HIV-1 infectivity is critically dependent on proper CA cone formation and mutations in CA are lethal when they inhibit CA assembly by destabilizing the intra and/or inter molecular CA contacts, which ultimately abrogate viral replication. Glu98, which is located on a surface of a flexible cyclophilin A binding loop is not involved in any intra-molecular contacts with other CA residues. In contrast, Glu187 has extensive intra-molecular contacts with eight other CA residues. Additionally, Glu187 has been shown to form a salt-bridge with Arg18 of another N-terminal CA monomer in a N-C dimer. However, despite proper virus release, glycoprotein incorporation and Gag processing, electron microscopy analysis revealed that, in contrast to the E187G mutant, only the E98A particles had aberrant core morphology that resulted in loss of infectivity.
| S-EPMC1832212 | biostudies-literature
Project description:The host cell factor cyclophilin A (CypA) interacts directly with the HIV-1 capsid and regulates viral infectivity. Although the crystal structure of CypA in complex with the N-terminal domain of the HIV-1 capsid protein (CA) has been known for nearly two decades, how CypA interacts with the viral capsid and modulates HIV-1 infectivity remains unclear. We determined the cryoEM structure of CypA in complex with the assembled HIV-1 capsid at 8-Å resolution. The structure exhibits a distinct CypA-binding pattern in which CypA selectively bridges the two CA hexamers along the direction of highest curvature. EM-guided all-atom molecular dynamics simulations and solid-state NMR further reveal that the CypA-binding pattern is achieved by single-CypA molecules simultaneously interacting with two CA subunits, in different hexamers, through a previously uncharacterized non-canonical interface. These results provide new insights into how CypA stabilizes the HIV-1 capsid and is recruited to facilitate HIV-1 infection.
| S-EPMC4785225 | biostudies-literature
Project description:Assembly of the HIV-1 virus involves, in part, strong interactions between the capsid (CA) domains of the Gag polyprotein. During maturation, the core of HIV-1 virions undergoes profound morphological changes due primarily to proteolysis of the CA domain from other Gag domains which may allow for more efficient disassembly of the viral core in the early stages of infection. The host protein cyclophilin A (CypA), a cis-trans prolyl isomerase, in some way seems to assist in this assembly/disassembly process. Using an unproteolyzed construct of CA, we show that binding of CypA induces a large-scale conformational change in CA that is independent of its cis-trans prolyl isomerase activity. This change appears to be mediated by Cys-198 of CA since mutation to Ala renders CypA unable to induce this change and alters the kinetics and stability of protein cores that may ultimately result in inefficient disassembly of viral cores. Alternately, mutation of the second CA Cys (C218A) allows for CypA-induced conformational changes but alters the kinetics and morphology of the protein cores that may ultimately result in inefficient assembly of viral cores. These studies show the importance of the CA Cys residues in mediating the contacts needed for viral assembly and disassembly.
| S-EPMC1305260 | biostudies-literature
Project description:Lentiviruses such as HIV-1 traverse nuclear pore complexes (NPC) and infect terminally differentiated non-dividing cells, but how they do this is unclear. The cytoplasmic NPC protein Nup358/RanBP2 was identified as an HIV-1 co-factor in previous studies. Here we report that HIV-1 capsid (CA) binds directly to the cyclophilin domain of Nup358/RanBP2. Fusion of the Nup358/RanBP2 cyclophilin (Cyp) domain to the tripartite motif of TRIM5 created a novel inhibitor of HIV-1 replication, consistent with an interaction in vivo. In contrast to CypA binding to HIV-1 CA, Nup358 binding is insensitive to inhibition with cyclosporine, allowing contributions from CypA and Nup358 to be distinguished. Inhibition of CypA reduced dependence on Nup358 and the nuclear basket protein Nup153, suggesting that CypA regulates the choice of the nuclear import machinery that is engaged by the virus. HIV-1 cyclophilin-binding mutants CA G89V and P90A favored integration in genomic regions with a higher density of transcription units and associated features than wild type virus. Integration preference of wild type virus in the presence of cyclosporine was similarly altered to regions of higher transcription density. In contrast, HIV-1 CA alterations in another patch on the capsid surface that render the virus less sensitive to Nup358 or TRN-SR2 depletion (CA N74D, N57A) resulted in integration in genomic regions sparse in transcription units. Both groups of CA mutants are impaired in replication in HeLa cells and human monocyte derived macrophages. Our findings link HIV-1 engagement of cyclophilins with both integration targeting and replication efficiency and provide insight into the conservation of viral cyclophilin recruitment.
| S-EPMC3234246 | biostudies-literature
Project description:The binding of cyclophilin A (CypA) to the human immunodeficiency virus type 1 (HIV-1) capsid protein (CA protein) is required soon after virus entry into natural target cells. In Jurkat T lymphocytes, disrupting CypA-CA interaction either by cyclosporine (Cs) treatment or by alteration (e.g., P90A) of the CA inhibits HIV-1 infection. In HeLa cells, however, treatment with Cs or Cs analogues minimally inhibits the early phase of HIV-1 infection but selects for a Cs-dependent virus with a change (A92E) in CA. To understand these phenomena, we examined the effects of the P90A and A92E changes in the HIV-1 CA protein on the stability of capsid complexes assembled in vitro and on capsid disassembly in the cytosol of virus-exposed target cells. The A92E change impaired CA-CA interactions in vitro and decreased the amount of particulate capsids in the cytosol of HeLa target cells. Reducing the binding of CypA to the A92E mutant capsid, either by Cs treatment or by an additional P90A change in the CA protein, increased the amount of particulate capsids and viral infectivity in HeLa cells. In contrast, reduction of the binding of CypA to HIV-1 capsids in Jurkat T lymphocytes resulted in a decrease in the amount of particulate capsids and infectivity. Thus, depending on the capsid and the target cell, CypA-CA binding either stabilized or destabilized the capsid, indicating that CypA modulates HIV-1 capsid disassembly. In both cell types examined, decreased stability of the capsid was associated with a decrease in the efficiency of HIV-1 infection.
| S-EPMC2772774 | biostudies-literature