scholarly journals Structural insights into the conformational plasticity of the full-length trimeric HIV-1 envelope glycoprotein precursor

2018 ◽  
Author(s):  
Shijian Zhang ◽  
Wei Li Wang ◽  
Shuobing Chen ◽  
Maolin Lu ◽  
Eden P. Go ◽  
...  
Keyword(s):  
Viral Entry ◽  
Envelope Glycoprotein ◽  
Structural Data ◽  
Full Length ◽  
Heptad Repeat ◽  
Structural Basis ◽  
Glycoprotein Precursor ◽  
Helix Bundle ◽  
Conformational Plasticity ◽  
Hiv 1

SummaryThe human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer mediates viral entry into cells and is the major target for the host antibody response. In infected cells, the mature Env [(gp120/gp41)3] is produced by cleavage of a trimeric gp160 precursor. Proteolytic cleavage decreases Env conformational flexibility, allowing the mature Env to resist antibody binding to conserved elements. The conformational plasticity of the Env precursor skews the humoral immune response towards the elicitation of ineffectual antibodies, contributing to HIV-1 persistence in the infected host. The structural basis for the plasticity of the Env precursor remains elusive. Here we use cryo-electron microscopy to visualize two coexisting conformational states of the full-length Env precursor at nominal resolutions of 5.5 and 8.0 Å. The State-P2 conformation features a three-helix bundle of the gp41 heptad repeat region in the core, but has disordered membrane-interactive regions. State-P1 trimers lack the three-helix bundle and instead retain ordered transmembrane and membrane-proximal external regions embracing a central cavity. Our structural data shed light on the unusual plasticity of the Env precursor and provide new clues to Env immunogen discovery.

scholarly journals Conserved Salt Bridge between the N- and C-Terminal Heptad Repeat Regions of the Human Immunodeficiency Virus Type 1 gp41 Core Structure Is Critical for Virus Entry and Inhibition

2008 ◽  
Vol 82 (22) ◽  
pp. 11129-11139 ◽  
Author(s):  
Yuxian He ◽  
Shuwen Liu ◽  
Jingjing Li ◽  
Hong Lu ◽  
Zhi Qi ◽  
...  
Keyword(s):  
Viral Entry ◽  
Salt Bridge ◽  
Core Structure ◽  
Heptad Repeat ◽  
Helix Bundle ◽  
Fusion Inhibitors ◽  
Immunodeficiency Virus ◽  
Hiv 1 ◽  
Positively Charged

ABSTRACT The fusogenic human immunodeficiency virus type 1 (HIV-1) gp41 core structure is a stable six-helix bundle formed by its N- and C-terminal heptad repeat sequences. Notably, the negatively charged residue Asp632 located at the pocket-binding motif in the C-terminal heptad repeat interacts with the positively charged residue Lys574 in the pocket formation region of the N-terminal heptad repeat to form a salt bridge. We previously demonstrated that the residue Lys574 plays an essential role in six-helix bundle formation and virus infectivity and is a key determinant of the target for anti-HIV fusion inhibitors. In this study, the functionality of residue Asp632 has been specifically characterized by mutational analysis and biophysical approaches. We show that Asp632 substitutions with positively charged residues (D632K and D632R) or a hydrophobic residue (D632V) could completely abolish Env-mediated viral entry, while a protein with a conserved substitution (D632E) retained its activity. Similar to the Lys574 mutations, nonconserved substitutions of Asp632 also severely impaired the α-helicity, stability, and conformation of six-helix bundles as shown by N36 and C34 peptides as a model system. Furthermore, nonconserved substitutions of Asp632 significantly reduced the potency of C34 to sequestrate six-helix bundle formation and to inhibit HIV-1-mediated cell-cell fusion and infection, suggesting its importance for designing antiviral fusion inhibitors. Taken together, these data suggest that the salt bridge between the N- and C-terminal heptad repeat regions of the fusion-active HIV-1 gp41 core structure is critical for viral entry and inhibition.

scholarly journals Mutations That Increase the Stability of the Postfusion gp41 Conformation of the HIV-1 Envelope Glycoprotein Are Selected by both an X4 and R5 HIV-1 Virus To Escape Fusion Inhibitors Corresponding to Heptad Repeat 1 of gp41, but the gp120 Adaptive Mutations Differ between the Two Viruses

2019 ◽  
Vol 93 (11) ◽  
Author(s):  
Min Zhuang ◽  
Russell Vassell ◽  
Chen Yuan ◽  
Paul W. Keller ◽  
Hong Ling ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Envelope Glycoprotein ◽  
Resistance Mutation ◽  
Heptad Repeat ◽  
Target Cells ◽  
Resistance Mutations ◽  
Adaptive Mutations ◽  
Helix Bundle ◽  
Fusion Inhibitors ◽  
Hiv 1

ABSTRACT Binding of the gp120 surface subunit of the envelope glycoprotein (Env) of HIV-1 to CD4 and chemokine receptors on target cells triggers refolding of the gp41 transmembrane subunit into a six-helix bundle (6HB) that promotes fusion between virus and host cell membranes. To elucidate details of Env entry and potential differences between viruses that use CXCR4 (X4) or CCR5 (R5) coreceptors, we generated viruses that are resistant to peptide fusion inhibitors corresponding to the first heptad repeat region (HR1) of gp41 that target fusion-intermediate conformations of Env. Previously we reported that an R5 virus selected two resistance pathways, each defined by an early gp41 resistance mutation in either HR1 or the second heptad repeat (HR2), to escape inhibition by an HR1 peptide, but preferentially selected the HR1 pathway to escape inhibition by a trimer-stabilized HR1 peptide. Here, we report that an X4 virus selected the same HR1 and HR2 resistance pathways as the R5 virus to escape inhibition by the HR1 peptide. However, in contrast to the R5 virus, the X4 virus selected a unique mutation in HR2 to escape inhibition by the trimer-stabilized peptide. Significantly, both of these X4 and R5 viruses acquired gp41 resistance mutations that improved the thermostability of the six-helix bundle, but they selected different gp120 adaptive mutations. These findings show that these X4 and R5 viruses use a similar resistance mechanism to escape from HR1 peptide inhibition but different gp120-gp41 interactions to regulate Env conformational changes. IMPORTANCE HIV-1 fuses with cells when the gp41 subunit of Env refolds into a 6HB after binding to cellular receptors. Peptides corresponding to HR1 or HR2 interrupt gp41 refolding and inhibit HIV infection. Previously, we found that a CCR5 coreceptor-tropic HIV-1 acquired a key HR1 or HR2 resistance mutation to escape HR1 peptide inhibitors but only the key HR1 mutation to escape a trimer-stabilized HR1 peptide inhibitor. Here, we report that a CXCR4 coreceptor-tropic HIV-1 selected the same key HR1 or HR2 mutations to escape inhibition by the HR1 peptide but different combinations of HR1 and HR2 mutations to escape the trimer-stabilized HR1 peptide. All gp41 mutations enhance 6HB stability to outcompete inhibitors, but gp120 adaptive mutations differed between these R5 and X4 viruses, providing new insights into gp120-gp41 functional interactions affecting Env refolding during HIV entry.

scholarly journals Structural Basis for Dodecameric Assembly States and Conformational Plasticity of the Full-Length AAA+ ATPases Rvb1·Rvb2

Structure ◽  
2015 ◽  
Vol 23 (3) ◽  
pp. 483-495 ◽  
Author(s):  
Kristina Lakomek ◽  
Gabriele Stoehr ◽  
Alessandro Tosi ◽  
Monika Schmailzl ◽  
Karl-Peter Hopfner
Keyword(s):  
Full Length ◽  
Structural Basis ◽  
Conformational Plasticity ◽  
Aaa Atpases

scholarly journals Similarities and differences between native HIV-1 envelope glycoprotein trimers and stabilized soluble trimer mimetics

2018 ◽  
Author(s):  
Alba Torrents de la Peña ◽  
Kimmo Rantalainen ◽  
Christopher A. Cottrell ◽  
Joel D. Allen ◽  
Marit J. van Gils ◽  
...  
Keyword(s):  
Envelope Glycoprotein ◽  
Neutralizing Antibodies ◽  
Transmembrane Domain ◽  
Cytoplasmic Tail ◽  
Full Length ◽  
Broadly Neutralizing Antibodies ◽  
Vaccine Research ◽  
Similarities And Differences ◽  
Membrane Tether ◽  
Hiv 1

AbstractThe HIV-1 envelope glycoprotein (Env) trimer is located on the surface of the virus and is the target of broadly neutralizing antibodies (bNAbs). Recombinant native-like soluble Env trimer mimetics, such as SOSIP trimers, have taken a central role in HIV-1 vaccine research aimed at inducing bNAbs. We therefore performed a direct and thorough comparison of a full-length native Env trimer containing the transmembrane domain and the cytoplasmic tail, with the sequence matched soluble SOSIP trimer, both based on an early Env sequence (AMC011) from an HIV+ individual that developed bNAbs. The structures of the full-length AMC011 trimer bound to either bNAb PGT145 or PGT151 were very similar to the structures of SOSIP trimers. Antigenically, the full-length and SOSIP trimers were comparable, but in contrast to the full-length trimer, the SOSIP trimer did not bind at all to non-neutralizing antibodies, most likely as a consequence of the intrinsic stabilization of the SOSIP trimer. Furthermore, the glycan composition of full-length and SOSIP trimers was similar overall, but the SOSIP trimer possessed slightly less complex and less extensively processed glycans, which may relate to the intrinsic stabilization as well as the absence of the membrane tether. These data provide insights into how to best use and improve membrane-associated full-length and soluble SOSIP HIV-1 Env trimers as immunogens.

scholarly journals Probing the Structure of the HIV-1 Envelope Trimer Using Aspartate Scanning Mutagenesis

2020 ◽  
Vol 94 (21) ◽  
Author(s):  
Raksha Das ◽  
Rohini Datta ◽  
Raghavan Varadarajan
Keyword(s):  
Cell Surface ◽  
Viral Entry ◽  
Fusion Peptide ◽  
Heptad Repeat ◽  
Subtype B ◽  
Clade C ◽  
Viral Surface ◽  
Virion Surface ◽  
Peptide Region ◽  
Hiv 1

ABSTRACT HIV-1 envelope (Env) glycoprotein gp160 exists as a trimer of heterodimers on the viral surface. In most structures of the soluble ectodomain of trimeric HIV-1 envelope glycoprotein, the regions from 512 to 517 of the fusion peptide and from 547 to 568 of the N-heptad repeat are disordered. We used aspartate scanning mutagenesis of subtype B strain JRFL Env as an alternate method to probe residue burial in the context of cleaved, cell surface-expressed Env, as buried residues should be intolerant to substitution with Asp. The data are inconsistent with a fully disordered 547 to 568 stretch, as residues 548, 549, 550, 555, 556, 559, 562, and 566 to 569 are all sensitive to Asp substitution. In the fusion peptide region, residues 513 and 515 were also sensitive to Asp substitution, suggesting that the fusion peptide may not be fully exposed in native Env. gp41 is metastable in the context of native trimer. Introduction of Asp at residues that are exposed in the prefusion state but buried in the postfusion state is expected to destabilize the postfusion state and any intermediate states where the residue is buried. We therefore performed soluble CD4 (sCD4)-induced gp120 shedding experiments to identify Asp mutants at residues 551, 554 to 559, 561 to 567, and 569 that could prevent gp120 shedding. We also observed similar mutational effects on shedding for equivalent mutants in the context of clade C Env from isolate 4-2J.41. These substitutions can potentially be used to stabilize native-like trimer derivatives that are used as HIV-1 vaccine immunogens. IMPORTANCE In most crystal structures of the soluble ectodomain of the HIV-1 Env trimer, some residues in the fusion and N-heptad repeat regions are disordered. Whether this is true in the context of native, functional Env on the virion surface is not known. This knowledge may be useful for stabilizing Env in its prefusion conformation and will also help to improve understanding of the viral entry process. Burial of the charged residue Asp in a protein structure is highly destabilizing. We therefore used Asp scanning mutagenesis to probe the burial of apparently disordered residues in native Env and to examine the effect of mutations in these regions on Env stability and conformation as probed by antibody binding to cell surface-expressed Env, CD4-induced shedding of HIV-1 gp120, and viral infectivity studies. Mutations that prevent shedding can potentially be used to stabilize native-like Env constructs for use as vaccine immunogens.

scholarly journals Understanding the structural basis of HIV-1 restriction by the full length double-domain APOBEC3G

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Hanjing Yang ◽  
Fumiaki Ito ◽  
Aaron D. Wolfe ◽  
Shuxing Li ◽  
Nazanin Mohammadzadeh ◽  
...  
Keyword(s):  
Full Length ◽  
Structural Basis ◽  
Hiv 1

scholarly journals Proteolytic Processing of the Human Immunodeficiency Virus Envelope Glycoprotein Precursor Decreases Conformational Flexibility

2012 ◽  
Vol 87 (3) ◽  
pp. 1884-1889 ◽  
Author(s):  
Hillel Haim ◽  
Ignacio Salas ◽  
Joseph Sodroski
Keyword(s):  
Human Immunodeficiency Virus ◽  
Envelope Glycoprotein ◽  
Conformational Flexibility ◽  
Proteolytic Processing ◽  
Virus Envelope ◽  
Glycoprotein Precursor ◽  
Trimeric Complex ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACTThe mature envelope glycoprotein (Env) spike on the surface of human immunodeficiency virus type 1 (HIV-1) virions is derived by proteolytic cleavage of a trimeric gp160 glycoprotein precursor. Remarkably, proteolytic processing of the HIV-1 Env precursor results in changes in Env antigenicity that resemble those associated with glutaraldehyde fixation. Apparently, proteolytic processing of the HIV-1 Env precursor decreases conformational flexibility of the Env trimeric complex, differentially affecting the integrity/accessibility of epitopes for neutralizing and nonneutralizing antibodies.

scholarly journals Structural Basis of CD4 Downregulation by HIV-1 Nef

2020 ◽  
Author(s):  
Yonghwa Kwon ◽  
Robyn Kaake ◽  
Ignacia Echeverria ◽  
Marissa Suarez ◽  
Charlotte Stoneham ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Viral Entry ◽  
Neutralizing Antibodies ◽  
Immune Surveillance ◽  
Underlying Mechanism ◽  
Host Cells ◽  
Structural Basis ◽  
Viral Glycoprotein ◽  
Class I Mhc ◽  
Hiv 1

The HIV-1 protein Nef suppresses multiple immune surveillance mechanisms to promote viral pathogenesis1. Individuals infected with HIV-1 encoding defective nef genes do not develop AIDS for decades2,3. A key target of Nef is the cellular receptor CD4. Although essential for viral entry into host cells, CD4 is problematic for the virus later in its replication cycle: CD4 disrupts processing of the viral glycoprotein, Env, inhibiting infectivity4; it interferes with the release of new virions5,6; and it causes vulnerability to superinfection, causing premature cell death and limiting viral productivity7. Furthermore, binding of CD4 to Env exposes otherwise-concealed Env epitopes, rendering infected cells more susceptible to antibody-dependent cellular cytotoxicity and virus particles more susceptible to neutralizing antibodies8-10. HIV-1 has evolved strategies to mitigate these problems. Newly synthesized CD4 is targeted in the endoplasmic reticulum by the viral Vpu protein for proteasomal degradation11. Surface-expressed CD4, in contrast, is targeted by Nef for endocytosis and lysosomal degradation12-15. Nef’s effect on CD4 involves hijacking of clathrin adaptor complex 2 (AP2)-dependent endocytosis16,17. Although how Nef associates with a part of the tetrameric AP2 is understood18, a complete understanding of the interaction, especially how CD4 is sequestered by Nef into a complex with AP2, has remained elusive. Here, we present a high-resolution crystal structure that describes the underlying mechanism. An intricate combination of conformational changes occurs in both Nef and AP2 to enable CD4 binding and downregulation. Strikingly, a pocket on Nef previously identified as crucial for recruiting class I MHC is also responsible for recruiting CD4, revealing a potential approach to inhibit two of Nef’s activities and sensitize the virus to immune clearance

scholarly journals Targeted destabilization of the HIV-1 gp120-gp41 interface leads to convergent evolution with mutations in the V1V2, HR1 and HR2 domains

2021 ◽  
Author(s):  
Alba Torrents de la Peña ◽  
Iván del Moral Sánchez ◽  
Judith A. Burger ◽  
Ilja Bontjer ◽  
Gözde Isik ◽  
...  
Keyword(s):  
Viral Entry ◽  
Envelope Glycoprotein ◽  
Convergent Evolution ◽  
High Throughput Screening ◽  
Neutralizing Antibodies ◽  
Dominant Role ◽  
Virus Evolution ◽  
Target Cells ◽  
Compensatory Mutations ◽  
Hiv 1

The HIV-1 envelope glycoprotein (Env) trimer is responsible for viral entry into target cells and is the sole target of neutralizing antibodies. The Env protein is therefore the focus of HIV-1 vaccine design. Env consists of two non-covalently linked subunits (gp120 and gp41) that form a trimer of heterodimers and this 6-subunit complex is metastable and conformationally flexible. Several approaches have been pursued to stabilize the Env trimer for vaccine purposes, which include structure-based design, high-throughput screening and selection by mammalian cell display. Here, we employed directed virus evolution to improve Env trimer stability. Accordingly, we deliberately destabilized the Env gp120-gp41 interface by mutagenesis in the context of replicating HIV-1 LAI virus and virus evolution over time. We identified compensatory changes that pointed at convergent evolution as they were largely restricted to specific Env regions, namely the V1V2-domain of gp120, and the the HR1 and HR2 domain of gp41. Specifically, S614G in V1V2 and Q567R in HR1 were frequently identified. Interestingly, the majority of the compensatory mutations were at distant locations from the original mutations and most likely strengthen inter-subunit interactions. These results show how the virus can overcome Env instability and illuminate the regions that play a dominant role in Env stability. Importance A successful HIV-1 vaccine most likely requires an envelope glycoprotein (Env) component, as the Env is the only viral protein on the surface of the virus and the target for neutralizing antibodies. However, HIV Env is metastable and flexible because of the weak interactions between the Env subunits, complicating the generation of recombinant mimics of native Env. Here, we used directed viral evolution to study Env stability. We deliberately destabilized the interface between Env subunits and explored the capacity of the virus to repair trimer instability by evolution. We identified compensatory mutations that converged in specific Env locations: the apex and the trimer interface. Selected mutations enhanced the stability of recombinant soluble Env trimer proteins. These results provided clues on understanding the structural mechanisms involved in Env trimer stability, which can guide future immunogen design.

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