Crystal Structure of the Ebola Virus Membrane Fusion Subunit, GP2, from the Envelope Glycoprotein Ectodomain

Weissenhorn, Winfríed; Carfı́, Andrea; Lee, Kon-Ho; Skehel, J.J.; Wiley, Don C.

doi:10.1016/s1097-2765(00)80159-8

Cited by 383 publications

(435 citation statements)

References 73 publications

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“…The HR2 helix is oblique with respect to the HR1 trimer axis and does not form a classical coiled-coil-like interaction with the HR1 helices, as observed for the C͞N peptide packing of HIV-1 gp41 (30,31) and Ebola Gp2 (33,34). The interaction with the HR1 trimer is mainly hydrophobic, with apolar amino acids of the HR2 helix (Ile-1161, Leu-1168, Val-1171, and Leu-1175) packing against the HR1 trimer grooves and only three interactions formed by charged͞polar residues.…”

Section: Resultsmentioning

confidence: 82%

Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

Supekar¹,

Bruckmann²,

Ingallinella³

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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116

111

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A coronavirus (CoV) has recently been identified as the causative agent of the severe acute respiratory syndrome (SARS) in humans. CoVs enter target cells through fusion of viral and cellular membranes mediated by the viral envelope glycoprotein S. We have determined by x-ray crystallography the structure of a proteolytically stable core fragment from the heptad repeat (HR) regions HR1 and HR2 of the SARS-CoV S protein. We have also determined the structure of an HR1-HR2 S core fragment, containing a shorter HR1 peptide and a C-terminally longer HR2 peptide that extends up to the transmembrane region. In these structures, three HR1 helices form a parallel coiled-coil trimer, whereas three HR2 peptides pack in an oblique and antiparallel fashion into the coiled-coil hydrophobic grooves, adopting mixed extended and ␣-helical conformations as in postfusion paramyxoviruses F proteins structures. coiled-coil ͉ membrane fusion ͉ viral entry ͉ heptad repeat

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Section: Resultsmentioning

confidence: 82%

Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

Supekar¹,

Bruckmann²,

Ingallinella³

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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116

111

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“…The prehairpin intermediate resolves into the trimer-ofhairpins structure, resulting in membrane fusion. The trimer-of-hairpins motif has been observed in several crystal structures of viral envelope proteins [2][3][4][5][6][7][8][9][10][11][12][13][14][15] . In this motif, the N-peptides fold into a central parallel homotrimeric coiled coil, with three C-peptides packing in an antiparallel manner along the conserved hydrophobic grooves on the coiled-coil surface.…”

Section: Resultsmentioning

confidence: 99%

“…Asparagine, glutamine, and threonine residues are commonly observed in the nonpolar cores of viral coiled coils (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15). Interestingly, the Visna VN40͞VC34 coiled-coil core contains serine residues in position d of the heptad repeat.…”

Section: Resultsmentioning

confidence: 99%

“…In this motif, a homotrimeric coiled coil, formed by the N-terminal regions of the protein, is surrounded by three C-terminal regions that pack against the coiled coil in an oblique antiparallel manner. In a large number of known viral protein structures, including influenza hemagglutinin (2, 3), human and simian immunodeficiency virus gp41 (4-9), Moloney murine leukemia virus TM (10), Ebola virus GP2 (11,12), human T-cell leukemia virus type 1 gp21 (13), simian parainfluenza virus 5 F1 (14), human respiratory syncytial virus F1 (15), and Newcastle disease virus F (16), the trimer-of-hairpins motif ensures that the fusion peptide (which is N-terminal of the coiled-coil domain) is spatially close to the transmembrane segment at the C terminus ( Figs. 1 and 2).…”

mentioning

confidence: 99%

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The trimer-of-hairpins motif in membrane fusion: Visna virus

Malashkevich

Singh²,

Kim³

2001

Proc. Natl. Acad. Sci. U.S.A.

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Structural studies of viral membrane fusion proteins suggest that a ''trimer-of-hairpins'' motif plays a critical role in the membrane fusion process of many enveloped viruses. In this motif, a coiled coil (formed by homotrimeric association of the N-terminal regions of the protein) is surrounded by three C-terminal regions that pack against the coiled coil in an oblique antiparallel manner. The resulting trimer-of-hairpins structure serves to bring the viral and cellular membranes together for fusion. LEARNCOIL-VMF, a computational program developed to recognize coiled coil-like regions that form the trimer-of-hairpins motif, predicts these regions in the membrane fusion protein of the Visna virus. Peptides corresponding to the computationally identified sequences were synthesized, and the soluble core of the Visna membrane fusion protein was reconstituted in solution. Its crystal structure at 1.5-Å resolution demonstrates that a trimer-of-hairpins structure is formed. Remarkably, despite less than 23% sequence identity, the ectodomains in Visna and HIV-1 envelope glycoproteins show detailed structural conservation, especially within the area of a hydrophobic pocket in the central coiled coil currently being targeted for the development of new anti-HIV drugs.S urface glycoproteins of enveloped viruses play a critical role in viral infectivity and replication (1). These envelope proteins mediate the initial virion attachment to the cell and the subsequent fusion of the viral and cellular membranes (Fig. 1). An emerging consensus from structural studies of viral membrane fusion proteins suggests that a large number of diverse enveloped viruses use the ''trimer-of-hairpins'' motif for viral entry. In this motif, a homotrimeric coiled coil, formed by the N-terminal regions of the protein, is surrounded by three C-terminal regions that pack against the coiled coil in an oblique antiparallel manner. In a large number of known viral protein structures, including influenza hemagglutinin (2, 3), human and simian immunodeficiency virus gp41 (4-9), Moloney murine leukemia virus TM (10), Ebola virus GP2 (11, 12), human T-cell leukemia virus type 1 gp21 (13), simian parainfluenza virus 5 F1 (14), human respiratory syncytial virus F1 (15), and Newcastle disease virus F (16), the trimer-of-hairpins motif ensures that the fusion peptide (which is N-terminal of the coiled-coil domain) is spatially close to the transmembrane segment at the C terminus ( Figs. 1 and 2). Given that the fusion peptide inserts into the cell membrane and the transmembrane segment is anchored in the viral membrane, formation of the trimer-of-hairpins motif is thought to facilitate the apposition of the viral and cellular membranes. But whether hairpin formation precedes the actual membrane fusion event or occurs simultaneously with fusion is unknown (see Fig. 1 legend).In earlier work we developed a specialized program, LEARN-COIL-VMF, for identifying potential coiled coil-like regions that make up the trimer-of-hairpins motif in viral membrane fusion ...

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“…These fusion peptides are flanked by two Cys residues, which, based on structural and mutagenesis evidence, likely form a disulfide bond (9,21). The ASLV fusion peptide contains one proline, and the filovirus fusion peptides contain two prolines, at their centers.…”

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confidence: 99%

Cysteines Flanking the Internal Fusion Peptide Are Required for the Avian Sarcoma/Leukosis Virus Glycoprotein To Mediate the Lipid Mixing Stage of Fusion with High Efficiency

Delos¹,

Brecher

Chen³

et al. 2008

J Virol

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We previously showed that the cysteines flanking the internal fusion peptide of the avian sarcoma/leukosis virus subtype A (ASLV-A) Env (EnvA) are important for infectivity and cell-cell fusion. Here we define the stage of fusion at which the cysteines are required. The flanking cysteines are dispensable for receptor-triggered membrane association but are required for the lipid mixing step of fusion, which, interestingly, displays a high pH onset and a biphasic profile. Second-site mutations that partially restore infection partially restore lipid mixing. These findings indicate that the cysteines flanking the internal fusion peptide of EnvA (and perhaps by analogy Ebola virus glycoprotein) are important for the foldback stage of the conformational changes that lead to membrane merger.The avian sarcoma/leukosis virus (ASLV) Env protein is unusual among class I fusion proteins in that it contains an internal fusion peptide, a characteristic it shares with filovirus (Ebola virus and Marburg virus) glycoproteins. These fusion peptides are flanked by two Cys residues, which, based on structural and mutagenesis evidence, likely form a disulfide bond (9, 21). The ASLV fusion peptide contains one proline, and the filovirus fusion peptides contain two prolines, at their centers. Previous work has shown that these Pro residues are important for fusion (4,8). The fusion peptide of EnvA is operationally defined as residues 22 to 37 of the transmembrane (TM) subunit. We previously provided evidence that the Pro in the middle of this peptide (P29) requires a ␤-turn structure (4) at some stage of fusion. We subsequently showed that the two Cys residues (C9 and C45) that define the likely disulfide-bonded loop encompassing the fusion peptide are required for infectivity and cell-cell fusion (5). In this study, we identified the specific stage of fusion that requires the Cys residues that flank the EnvA fusion peptide.To begin our studies, we asked if the defect was in the first step of fusion: target membrane binding. Accordingly, we determined the ability of murine leukemia virus pseudotyped virions (10) bearing either wild-type EnvA or an EnvA harboring double Cys-to-Ser mutations at both positions 9 and 45 of the TM subunit (referred to below as EnvAC9,45S) to bind to target membranes in a receptor-dependent manner (6, 7, 15). As seen in Fig. 1, wild-type murine leukemia virus pseudovirions bound liposomes and floated to the top of the gradient when either the quail (lane 1) or the chicken (lane 5) form of the soluble Tva receptor (sTva) was used (7). In the absence of sTva, all of the pseudovirions remained at the bottom of the gradient (Fig. 1, lane 12). Similar receptor-dependent membrane association was observed for EnvAC9,45S (Fig. 1). These data show that the Cys residues (C9 and C45) that define the loop encompassing the fusion peptide are not required for receptor-triggered binding of EnvA-bearing virus particles to target membranes.We then asked whether the cysteines are required for EnvA to reach the lipid mixing ...

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Crystal Structure of the Ebola Virus Membrane Fusion Subunit, GP2, from the Envelope Glycoprotein Ectodomain

Cited by 383 publications

References 73 publications

Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

Structure of a proteolytically resistant core from the severe acute respiratory syndrome coronavirus S2 fusion protein

The trimer-of-hairpins motif in membrane fusion: Visna virus

Cysteines Flanking the Internal Fusion Peptide Are Required for the Avian Sarcoma/Leukosis Virus Glycoprotein To Mediate the Lipid Mixing Stage of Fusion with High Efficiency

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