FRINK Linked Data Fragments server

Matches in SemOpenAlex for { <https://semopenalex.org/work/W2887352390> ?p ?o ?g. }

• W2887352390 endingPage "246.e4" @default.
• W2887352390 startingPage "235" @default.
• W2887352390 abstract "•SOSIP Env apex is 3-fold symmetric and consistent with closed prefusion structures•Unliganded Env base and CD4-bound Env apex and base exhibit flexibility•SOSIPs retain desired properties of immunogens; e.g., burying non-neutralizing epitopes•Results allow interpretation of smFRET studies and SOSIP and virion Env structures HIV-1 Envelope (Env) mediates viral-host membrane fusion after binding host-receptor CD4 and coreceptor. Soluble envelopes (SOSIPs), designed to mimic prefusion conformational states of virion-bound envelopes, are proposed immunogens for eliciting neutralizing antibodies, yet only static structures are available. To evaluate conformational landscapes of ligand-free, CD4-bound, inhibitor-bound, and antibody-bound SOSIPs, we measured inter-subunit distances throughout spin-labeled SOSIPs using double electron-electron resonance (DEER) spectroscopy and compared results to soluble and virion-bound Env structures, and single-molecule fluorescence resonance energy transfer (smFRET)-derived dynamics of virion-bound Envs. Unliganded SOSIP measurements were consistent with closed, neutralizing antibody-bound structures and shielding of non-neutralizing epitopes, demonstrating homogeneity at Env apex, increased flexibility near Env base, and no evidence for the intra-subunit flexibility near Env apex suggested by smFRET. CD4 binding increased inter-subunit distances and heterogeneity, consistent with rearrangements required for coreceptor binding. Results suggest similarities between SOSIPs and virion-bound Envs and demonstrate DEER’s relevance for immunogen design. HIV-1 Envelope (Env) mediates viral-host membrane fusion after binding host-receptor CD4 and coreceptor. Soluble envelopes (SOSIPs), designed to mimic prefusion conformational states of virion-bound envelopes, are proposed immunogens for eliciting neutralizing antibodies, yet only static structures are available. To evaluate conformational landscapes of ligand-free, CD4-bound, inhibitor-bound, and antibody-bound SOSIPs, we measured inter-subunit distances throughout spin-labeled SOSIPs using double electron-electron resonance (DEER) spectroscopy and compared results to soluble and virion-bound Env structures, and single-molecule fluorescence resonance energy transfer (smFRET)-derived dynamics of virion-bound Envs. Unliganded SOSIP measurements were consistent with closed, neutralizing antibody-bound structures and shielding of non-neutralizing epitopes, demonstrating homogeneity at Env apex, increased flexibility near Env base, and no evidence for the intra-subunit flexibility near Env apex suggested by smFRET. CD4 binding increased inter-subunit distances and heterogeneity, consistent with rearrangements required for coreceptor binding. Results suggest similarities between SOSIPs and virion-bound Envs and demonstrate DEER’s relevance for immunogen design. Developing a vaccine against HIV-1 requires understanding the structure and dynamics of envelope (Env) glycoproteins on virions and in soluble forms being developed as immunogens (Sanders and Moore, 2017Sanders R.W. Moore J.P. Native-like Env trimers as a platform for HIV-1 vaccine design.Immunol. Rev. 2017; 275: 161-182Crossref PubMed Scopus (167) Google Scholar). HIV-1 Env, a trimer of gp120-gp41 heterodimers, mediates entry into target cells by gp120 binding to the host receptor CD4, which initiates conformational changes that allow recognition of the coreceptor CCR5, resulting in gp41 rearrangements that promote fusion between the target cell and viral membranes (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar). Low-resolution reconstructions of Env trimers on HIV-1 virions derived by cryo-electron tomography (cryo-ET) revealed distinct Env conformations including an unliganded, closed structure in which adjacent gp120 subunits interacted to form the trimer apex and a soluble CD4 (sCD4)-bound, open conformation in which the gp120 subunits were displaced and outwardly rotated to disrupt the trimer apex (Liu et al., 2008Liu J. Bartesaghi A. Borgnia M.J. Sapiro G. Subramaniam S. Molecular architecture of native HIV-1 gp120 trimers.Nature. 2008; 455: 109-113Crossref PubMed Scopus (631) Google Scholar). Subsequent crystallographic and cryo-EM structures of soluble native-like Env trimers lacking membrane and cytoplasmic domains and including stabilizing mutations (SOSIPs) (Sanders et al., 2013Sanders R.W. Derking R. Cupo A. Julien J.P. Yasmeen A. de Val N. Kim H.J. Blattner C. de la Peña A.T. Korzun J. et al.A next-generation cleaved, soluble HIV-1 Env trimer, BG505 SOSIP.664 gp140, expresses multiple epitopes for broadly neutralizing but not non-neutralizing antibodies.PLoS Pathog. 2013; 9: e1003618Crossref PubMed Scopus (640) Google Scholar) in complex with broadly neutralizing antibodies (bNAbs) resulted in higher-resolution Env structures of the closed Env conformation, revealing interactions of the gp120 V1V2 motifs at the trimer apex that shield the coreceptor binding site on V3 (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar) (Figures 1A and 1B ). Consistent with cryo-ET structures of open virion-bound Envs (Liu et al., 2008Liu J. Bartesaghi A. Borgnia M.J. Sapiro G. Subramaniam S. Molecular architecture of native HIV-1 gp120 trimers.Nature. 2008; 455: 109-113Crossref PubMed Scopus (631) Google Scholar), single-particle cryo-EM structures of sCD4-bound SOSIPs demonstrated rotation and displacement of gp120s, an ∼40Å movement of V1V2 to the sides of Env trimer to reveal V3, and smaller rearrangements of gp41 (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar) (Figures 1A and 1B). The dynamics of HIV-1 Envs on virions has been characterized by single-molecule fluorescence resonance energy transfer (smFRET) studies, in which donor and acceptor fluorophores were placed in loops within V1 and V4 of a gp120 monomer in virion-bound Env trimers, allowing for intra-subunit motions within single Env trimers to be monitored over time (Ma et al., 2018Ma X. Lu M. Gorman J. Terry D.S. Hong X. Zhou Z. Zhao H. Altman R.B. Arthos J. Blanchard S.C. et al.HIV-1 Env trimer opens through an asymmetric intermediate in which individual protomers adopt distinct conformations.eLife. 2018; 7: 7Crossref Scopus (82) Google Scholar, Munro et al., 2014Munro J.B. Gorman J. Ma X. Zhou Z. Arthos J. Burton D.R. Koff W.C. Courter J.R. Smith 3rd, A.B. Kwong P.D. et al.Conformational dynamics of single HIV-1 envelope trimers on the surface of native virions.Science. 2014; 346: 759-763Crossref PubMed Scopus (339) Google Scholar, Munro and Lee, 2018Munro J.B. Lee K.K. Probing Structural Variation and Dynamics in the HIV-1 Env Fusion Glycoprotein.Curr. HIV Res. 2018; 16: 5-12Crossref PubMed Scopus (8) Google Scholar, Munro and Mothes, 2015Munro J.B. Mothes W. Structure and Dynamics of the Native HIV-1 Env Trimer.J. Virol. 2015; 89: 5752-5755Crossref PubMed Scopus (63) Google Scholar). These studies suggested that virion-bound Envs transition between three primary states (States 1, 2, and 3), with State 1 (low FRET ground state with large intra-dye distances) predominating in the absence of added ligands. Transitions to State 3 (medium FRET state with intermediate intra-dye distances) were induced by the addition of sCD4 plus the coreceptor-mimicking antibody 17b after populating State 2 (high FRET state with short intra-dye distances) (Ma et al., 2018Ma X. Lu M. Gorman J. Terry D.S. Hong X. Zhou Z. Zhao H. Altman R.B. Arthos J. Blanchard S.C. et al.HIV-1 Env trimer opens through an asymmetric intermediate in which individual protomers adopt distinct conformations.eLife. 2018; 7: 7Crossref Scopus (82) Google Scholar, Munro et al., 2014Munro J.B. Gorman J. Ma X. Zhou Z. Arthos J. Burton D.R. Koff W.C. Courter J.R. Smith 3rd, A.B. Kwong P.D. et al.Conformational dynamics of single HIV-1 envelope trimers on the surface of native virions.Science. 2014; 346: 759-763Crossref PubMed Scopus (339) Google Scholar). Addition of bNAbs or a small molecule inhibitor of HIV-1 entry, BMS-626529 (Li et al., 2013Li Z. Zhou N. Sun Y. Ray N. Lataillade M. Hanna G.J. Krystal M. Activity of the HIV-1 attachment inhibitor BMS-626529, the active component of the prodrug BMS-663068, against CD4-independent viruses and HIV-1 envelopes resistant to other entry inhibitors.Antimicrob. Agents Chemother. 2013; 57: 4172-4180Crossref PubMed Scopus (65) Google Scholar), stabilized the low-FRET ground state, with differential effects on the intermediate- and high-FRET states (Munro et al., 2014Munro J.B. Gorman J. Ma X. Zhou Z. Arthos J. Burton D.R. Koff W.C. Courter J.R. Smith 3rd, A.B. Kwong P.D. et al.Conformational dynamics of single HIV-1 envelope trimers on the surface of native virions.Science. 2014; 346: 759-763Crossref PubMed Scopus (339) Google Scholar). There is currently no correlation between atomic resolution Env structures and the smFRET-defined states, resulting in a gap in understanding the relationship between Env molecular structure and dynamics. In addition, the development of SOSIP Envs as potential immunogens (Sanders and Moore, 2017Sanders R.W. Moore J.P. Native-like Env trimers as a platform for HIV-1 vaccine design.Immunol. Rev. 2017; 275: 161-182Crossref PubMed Scopus (167) Google Scholar) requires assessment of whether their unliganded structures faithfully represent the conformation(s) adopted by virion-bound Envs that would be encountered in a natural infection. To evaluate the structure and conformational flexibility of SOSIP Envs in solution, we used double electron-electron resonance (DEER) spectroscopy to probe free and liganded SOSIP Envs spin-labeled with a nitroxide free radical. DEER measures the dipolar interaction between electron spin pairs and can be analyzed to give an interspin distance distribution in the range of 17–80Å (Jeschke, 2012Jeschke G. DEER distance measurements on proteins.Annu. Rev. Phys. Chem. 2012; 63: 419-446Crossref PubMed Scopus (722) Google Scholar). The populations of distances recorded reflect molecular motion in solution, providing a snapshot of protein dynamics and conformational heterogeneity. The largest peak amplitude defines the most probable distance in a DEER distribution and provides information on the dominant structural state in the population, while multiple peaks indicate conformational heterogeneity and peak widths are related to the internal flexibility of each conformation and the attached spin label (Hubbell et al., 2000Hubbell W.L. Cafiso D.S. Altenbach C. Identifying conformational changes with site-directed spin labeling.Nat. Struct. Biol. 2000; 7: 735-739Crossref PubMed Scopus (722) Google Scholar, Hubbell et al., 2013Hubbell W.L. López C.J. Altenbach C. Yang Z. Technological advances in site-directed spin labeling of proteins.Curr. Opin. Struct. Biol. 2013; 23: 725-733Crossref PubMed Scopus (224) Google Scholar). The accuracy of peak distance and width decrease as a function of interspin distance; 17–65Å distances can be assigned with confidence, whereas distances > 65Å are detected with less accuracy (e.g., +/−10Å) (Jeschke, 2012Jeschke G. DEER distance measurements on proteins.Annu. Rev. Phys. Chem. 2012; 63: 419-446Crossref PubMed Scopus (722) Google Scholar). Here we report DEER-derived inter-subunit distances and heterogeneity in unliganded SOSIP Envs and their complexes with sCD4, bNAbs, and a small molecule inhibitor. We also report an intra-subunit measurement between spin labels at positions similar to where dyes were introduced in smFRET studies of virion-bound Envs (Ma et al., 2018Ma X. Lu M. Gorman J. Terry D.S. Hong X. Zhou Z. Zhao H. Altman R.B. Arthos J. Blanchard S.C. et al.HIV-1 Env trimer opens through an asymmetric intermediate in which individual protomers adopt distinct conformations.eLife. 2018; 7: 7Crossref Scopus (82) Google Scholar, Munro et al., 2014Munro J.B. Gorman J. Ma X. Zhou Z. Arthos J. Burton D.R. Koff W.C. Courter J.R. Smith 3rd, A.B. Kwong P.D. et al.Conformational dynamics of single HIV-1 envelope trimers on the surface of native virions.Science. 2014; 346: 759-763Crossref PubMed Scopus (339) Google Scholar). These results inform models of Env dynamics relevant to developing SOSIP immunogens and to understanding conformational changes in Env-mediated membrane fusion. We chose to investigate two well-characterized SOSIP trimers, the clade A BG505 (Sanders et al., 2013Sanders R.W. Derking R. Cupo A. Julien J.P. Yasmeen A. de Val N. Kim H.J. Blattner C. de la Peña A.T. Korzun J. et al.A next-generation cleaved, soluble HIV-1 Env trimer, BG505 SOSIP.664 gp140, expresses multiple epitopes for broadly neutralizing but not non-neutralizing antibodies.PLoS Pathog. 2013; 9: e1003618Crossref PubMed Scopus (640) Google Scholar) and clade B B41 (Pugach et al., 2015Pugach P. Ozorowski G. Cupo A. Ringe R. Yasmeen A. de Val N. Derking R. Kim H.J. Korzun J. Golabek M. et al.A native-like SOSIP.664 trimer based on an HIV-1 subtype B env gene.J. Virol. 2015; 89: 3380-3395Crossref PubMed Scopus (187) Google Scholar), because these SOSIP Envs have been characterized structurally in closed and sCD4-bound open states (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar) and both are being developed as immunogens (Sanders et al., 2015Sanders R.W. van Gils M.J. Derking R. Sok D. Ketas T.J. Burger J.A. Ozorowski G. Cupo A. Simonich C. Goo L. et al.HIV-1 VACCINES. HIV-1 neutralizing antibodies induced by native-like envelope trimers.Science. 2015; 349: aac4223Crossref PubMed Scopus (383) Google Scholar). For each DEER experiment we used site-directed spin labeling (Hubbell et al., 2013Hubbell W.L. López C.J. Altenbach C. Yang Z. Technological advances in site-directed spin labeling of proteins.Curr. Opin. Struct. Biol. 2013; 23: 725-733Crossref PubMed Scopus (224) Google Scholar) to introduce a single cysteine into a gp120-gp41 protomer of the BG505 or B41 SOSIPs and then covalently attached a nitroxide spin label bearing the “V1” side chain (Khramtsov et al., 1989Khramtsov V.V. Yelinova V.I. Weiner L.M. Berezina T.A. Martin V.V. Volodarsky L.B. Quantitative determination of SH groups in low- and high-molecular-weight compounds by an electron spin resonance method.Anal. Biochem. 1989; 182: 58-63Crossref PubMed Scopus (74) Google Scholar) (indicated as an asterisk,∗) (Figure S1A). Once attached, the V1 spin label is about the size of an amino acid side chain and contributes limited width to DEER distance distributions (Toledo Warshaviak et al., 2013Toledo Warshaviak D. Khramtsov V.V. Cascio D. Altenbach C. Hubbell W.L. Structure and dynamics of an imidazoline nitroxide side chain with strongly hindered internal motion in proteins.J. Magn. Reson. 2013; 232: 53-61Crossref PubMed Scopus (31) Google Scholar) (Figures S1A and S1B). This approach results in the attachment of three spin labels per trimer, which form the vertices of an equilateral triangle if located in a symmetric portion of Env (Figure 1B), or the vertices of an isosceles or scalene triangle if located in a region that lacks 3-fold symmetry and/or adopts multiple conformations. Accordingly, DEER measurements were expected to result in one distance corresponding to the sides of an equilateral triangle or ≥ 2 distances corresponding to the sides of an asymmetric triangle or equilateral triangles of different sizes. We selected spin labeling sites from solvent-exposed residues in defined secondary structures (β strand or α-helix; not loops) in the following Env structural motifs in the apex or base regions: (1) (apex) gp120 V1V2, which mediates inter-subunit contacts that form the apex of closed Env trimers and repositions upon Env binding to sCD4 (Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar); (2) (apex) gp120 V3, which is shielded by V1V2 in the closed Env conformation and is exposed to provide coreceptor-binding sites upon Env binding to sCD4 (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar); (3) (apex) gp120 bridging sheet at the base of V1V2, which undergoes structural rearrangements upon CD4 binding that stabilize open conformations (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar); (4) (base) gp120 inner domain, which interacts with gp41 (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar); and (5) (base) gp41, portions of which undergo conformational changes upon binding to sCD4 (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar) (Figures 1A and 1B). To evaluate the positions of spin labeling sites among published structures, we measured inter-subunit Cα distances from crystal and cryo-EM structure coordinates and determined a mean distance and SD for each site (Figure 1C). Measurements included seven closed Envs (six SOSIPs) (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar) and one native (non-SOSIP) Env (Lee et al., 2016Lee J.H. Ozorowski G. Ward A.B. Cryo-EM structure of a native, fully glycosylated, cleaved HIV-1 envelope trimer.Science. 2016; 351: 1043-1048Crossref PubMed Scopus (315) Google Scholar), each complexed with one or more bNAb Fabs, B41 SOSIP bound to the CD4 binding site (CD4bs) bNAb b12 (B41-b12 complex) (Ozorowski et al., 2017), BG505 SOSIP bound to sCD4, the CD4-induced (CD4i) coreceptor-mimicking antibody 17b, and the gp120-gp41 interface bNAb 8ANC195 (BG505-sCD4-17b-8ANC195 complex) (Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar), and B41 SOSIP bound to sCD4 and 17b (B41-sCD4-17b complex) (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar). For closed Envs structures, the mean inter-subunit distance SD were ∼1Å or less, with the exception of residue 657 in gp41, whose position was different in two structures (pdb codes 5U70 and 5U7M), resulting in a 2.8Å SD (Figure 1C). We used these coordinate-derived measurements as references to compare with DEER-derived distances, noting that a coordinate-derived Cα-Cα measurement and an experimental DEER distance may differ by several Ångstrom yet represent the same structure because V1 side chain rotamers could contribute to the DEER distance (Figure S1B). To evaluate Env flexibility and heterogeneity in V1V2, which forms the closed trimer apex and is repositioned upon CD4 binding, we spin-labeled BG505 β strand residue 173 to make BG505-173∗. Distance distributions calculated from DEER spectra for unliganded BG505-173∗ revealed a most probable distance peak at 38Å, with a full width at half maximum (FWHM) of ∼10Å broadening to ≥ 15Å at its base (Figures 2A and S2), consistent with Cα-Cα measurements of closed Env structures (mean = 36Å) (Figure 1C) and suggesting 3-fold symmetry and detectable, but limited, structural heterogeneity. Unliganded B41-173∗ also exhibited a most probable distance at 38Å, similar to the BG505-173∗ peak, albeit with a reduced FWHM and a lower probability shoulder in the 29–35Å range, indicative of structural homogeneity in V1V2 (Figures 2D and S2). The addition of sCD4 to BG505-173∗ and B41-173∗ induced a large reduction of interspin pairs, suggesting that the majority of 173∗ spin labels moved beyond the ∼80Å limit of DEER detection, consistent with cryo-EM structures of sCD4-bound SOSIP Envs in which V1V2 is displaced to the sides of Env trimer and in which the three copies of residue 173 are separated by ∼110Å) (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar; H. Wang and P.J.B., unpublished data; Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar) (Figures 2A and 2D; S2). We also detected a small heterogeneous population (< 10% of total) of short (∼25Å) and long (> 65Å) interspin distance peaks for both samples (Figures 2A and 2D), which may reflect defined structures in a sCD4-bound conformation. We investigated the conformational flexibility of V3, the binding site for coreceptor, which is buried beneath V1V2 in closed structures and exposed upon CD4 binding (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar), by spin labeling BG505 residue 306 (Figure 1). DEER distributions for unliganded BG505-306∗ exhibited a narrow peak at 40Å with a < 5Å FWHM (Figure 2B), indicating that the unliganded BG505 Env structure is 3-fold symmetric and rigid with respect to V3. Upon binding to sCD4, a decrease in interspin signal indicated that some spin labels moved out of range, while a smaller population exhibited a peak at 65Å (Figure 2B). B41-306∗ and B41-306∗–sCD4 distance distributions yielded similar results, with a narrow distribution at 38Å for unliganded samples and increased heterogeneity characterized by a broad peak centered at 65Å upon the addition of sCD4 (Figure 2E). These results are consistent with closed and sCD4-bound Env structures: the inter-subunit distance separating residue 306 Cα atoms in closed bNAb-bound Env (Pancera et al., 2014Pancera M. Zhou T. Druz A. Georgiev I.S. Soto C. Gorman J. Huang J. Acharya P. Chuang G.Y. Ofek G. et al.Structure and immune recognition of trimeric pre-fusion HIV-1 Env.Nature. 2014; 514: 455-461Crossref PubMed Scopus (563) Google Scholar) is 37Å, and residue 306 is either disordered (BG505-sCD4-17b-8ANC195) or separated by 78Å (B41-sCD4-17b) in sCD4-bound Env structures (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar) (Figure 1C). The ∼13Å difference in inter-subunit distances between B41-306∗-sCD4 DEER measurements (65Å) and the B41-sCD4-17b cryo-EM structure (78Å) suggests that the B41-sCD4 complex can adopt shorter inter-subunit distances between V3 loops in the absence of 17b Fab, which may stabilize a larger inter-subunit separation though interactions near residue 306. We next labeled residue 202 in the gp120 β3 strand that becomes part of the bridging sheet upon sCD4 binding (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar). Distance distributions for unliganded BG505-202∗ revealed a most probable distance at 24Å with a ∼10Å FWHM (Figure 2C), consistent with closed bNAb-bound Env structures (Figure 1C). Adding sCD4 to BG505-202∗ broadened the FWHM to 20Å with an overall 50Å span; it also shifted most probable distances, resulting in peaks at 36Å and 42Å (Figure 2C). The B41-202∗ and BG505-202∗ distance distributions were essentially superimposable, and the B41-202∗–sCD4 distance distribution also showed that addition of sCD4 increased conformational heterogeneity, including a new peak at 44Å (Figure 2F). The emergence of long distance peaks upon sCD4 binding is consistent with residue 202 Cα atoms being separated by 48Å (BG505-sCD4-17b-8ANC195) or 64Å (B41-sCD4-17b) in sCD4-bound Env structures (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar) (Figure 1C), and the existence of multiple distances suggests the existence of sCD4-bound conformations that have not been observed in X-ray and cryo-EM Env structures. To study the gp120 inner domain, a region near the middle of Env trimer that contacts neighboring gp120 subunits and the gp41 subunit in the same protomer (Ward and Wilson, 2017Ward A.B. Wilson I.A. The HIV-1 envelope glycoprotein structure: nailing down a moving target.Immunol. Rev. 2017; 275: 21-32Crossref PubMed Scopus (169) Google Scholar), we spin-labeled gp120 residue 106. Unliganded BG505-106∗ distance distributions exhibited a 27Å most probable distance (Figure 2G), consistent with Cα-Cα distances measured in bNAb-bound BG505 crystal structures (30Å) (Figure 1C). We also observed a population of lower probability (∼25%) peaks at 35Å, 40Å, 50Å, and 61Å, indicating conformational heterogeneity in the inner domain of unliganded SOSIP Env (Figure 2G). The addition of sCD4 induced a distance distribution shift in which peaks at 40Å, 48Å, and 58Å were populated, with the 48Å peak being the most probable. These results are consistent with residue 106 Cα atoms being separated by 46Å (BG505-sCD4-17b-8ANC195) or 51Å (B41-sCD4-17b) in sCD4-bound Env structures (Ozorowski et al., 2017Ozorowski G. Pallesen J. de Val N. Lyumkis D. Cottrell C.A. Torres J.L. Copps J. Stanfield R.L. Cupo A. Pugach P. et al.Open and closed structures reveal allostery and pliability in the HIV-1 envelope spike.Nature. 2017; 547: 360-363Crossref PubMed Scopus (155) Google Scholar, Wang et al., 2016Wang H. Cohen A.A. Galimidi R.P. Gristick H.B. Jensen G.J. Bjorkman P.J. Cryo-EM structure of a CD4-bound open HIV-1 envelope trimer reveals structural rearrangements of the gp120 V1V2 loop.Proc. Natl. Acad. Sci. USA. 2016; 113: E7151-E7158Crossref PubMed Scopus (91) Google Scholar) (Figure 1C) and also suggest the presence of sCD4-bound conf" @default.
• W2887352390 created "2018-08-22" @default.
• W2887352390 creator A5036129613 @default.
• W2887352390 creator A5036133609 @default.
• W2887352390 creator A5062320657 @default.
• W2887352390 creator A5068857084 @default.
• W2887352390 creator A5070603489 @default.
• W2887352390 creator A5077255841 @default.
• W2887352390 creator A5088314450 @default.
• W2887352390 date "2018-08-01" @default.
• W2887352390 modified "2023-10-15" @default.
• W2887352390 title "DEER Spectroscopy Measurements Reveal Multiple Conformations of HIV-1 SOSIP Envelopes that Show Similarities with Envelopes on Native Virions" @default.
• W2887352390 cites W1858237937 @default.
• W2887352390 cites W1895742673 @default.
• W2887352390 cites W2008830902 @default.
• W2887352390 cites W2016180326 @default.
• W2887352390 cites W2022098078 @default.
• W2887352390 cites W2040595716 @default.
• W2887352390 cites W2044712738 @default.
• W2887352390 cites W2052708596 @default.
• W2887352390 cites W2062204035 @default.
• W2887352390 cites W2065241264 @default.
• W2887352390 cites W2074179519 @default.
• W2887352390 cites W2082355480 @default.
• W2887352390 cites W2083002978 @default.
• W2887352390 cites W2085577978 @default.
• W2887352390 cites W2092848363 @default.
• W2887352390 cites W2095224834 @default.
• W2887352390 cites W2098513990 @default.
• W2887352390 cites W2114227017 @default.
• W2887352390 cites W2118274231 @default.
• W2887352390 cites W2118820599 @default.
• W2887352390 cites W2120031187 @default.
• W2887352390 cites W2132629607 @default.
• W2887352390 cites W2142844453 @default.
• W2887352390 cites W2145002856 @default.
• W2887352390 cites W2147917005 @default.
• W2887352390 cites W2166730438 @default.
• W2887352390 cites W2197610643 @default.
• W2887352390 cites W2210879955 @default.
• W2887352390 cites W2268112994 @default.
• W2887352390 cites W2332658587 @default.
• W2887352390 cites W2336507460 @default.
• W2887352390 cites W2380132702 @default.
• W2887352390 cites W2515956381 @default.
• W2887352390 cites W2520562417 @default.
• W2887352390 cites W2521026695 @default.
• W2887352390 cites W2541922763 @default.
• W2887352390 cites W2581517068 @default.
• W2887352390 cites W2581941486 @default.
• W2887352390 cites W2606178393 @default.
• W2887352390 cites W2734881930 @default.
• W2887352390 cites W2750272252 @default.
• W2887352390 cites W2751727630 @default.
• W2887352390 cites W2781210171 @default.
• W2887352390 cites W2792975318 @default.
• W2887352390 doi "https://doi.org/10.1016/j.immuni.2018.06.017" @default.
• W2887352390 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/6104740" @default.
• W2887352390 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/30076100" @default.
• W2887352390 hasPublicationYear "2018" @default.
• W2887352390 type Work @default.
• W2887352390 sameAs 2887352390 @default.
• W2887352390 citedByCount "61" @default.
• W2887352390 countsByYear W28873523902018 @default.
• W2887352390 countsByYear W28873523902019 @default.
• W2887352390 countsByYear W28873523902020 @default.
• W2887352390 countsByYear W28873523902021 @default.
• W2887352390 countsByYear W28873523902022 @default.
• W2887352390 countsByYear W28873523902023 @default.
• W2887352390 crossrefType "journal-article" @default.
• W2887352390 hasAuthorship W2887352390A5036129613 @default.
• W2887352390 hasAuthorship W2887352390A5036133609 @default.
• W2887352390 hasAuthorship W2887352390A5062320657 @default.
• W2887352390 hasAuthorship W2887352390A5068857084 @default.
• W2887352390 hasAuthorship W2887352390A5070603489 @default.
• W2887352390 hasAuthorship W2887352390A5077255841 @default.
• W2887352390 hasAuthorship W2887352390A5088314450 @default.
• W2887352390 hasBestOaLocation W28873523901 @default.
• W2887352390 hasConcept C12554922 @default.
• W2887352390 hasConcept C159047783 @default.
• W2887352390 hasConcept C3013748606 @default.
• W2887352390 hasConcept C86803240 @default.
• W2887352390 hasConceptScore W2887352390C12554922 @default.
• W2887352390 hasConceptScore W2887352390C159047783 @default.
• W2887352390 hasConceptScore W2887352390C3013748606 @default.
• W2887352390 hasConceptScore W2887352390C86803240 @default.
• W2887352390 hasFunder F4320306137 @default.
• W2887352390 hasFunder F4320332161 @default.
• W2887352390 hasIssue "2" @default.
• W2887352390 hasLocation W28873523901 @default.
• W2887352390 hasLocation W28873523902 @default.
• W2887352390 hasLocation W28873523903 @default.
• W2887352390 hasLocation W28873523904 @default.
• W2887352390 hasLocation W28873523905 @default.
• W2887352390 hasOpenAccess W2887352390 @default.
• W2887352390 hasPrimaryLocation W28873523901 @default.
• W2887352390 hasRelatedWork W1702322095 @default.
• W2887352390 hasRelatedWork W1865764402 @default.

• next