scholarly journals Viral fusion protein transmembrane domain adopts β-strand structure to facilitate membrane topological changes for virus–cell fusion

2015 ◽  
Vol 112 (35) ◽  
pp. 10926-10931 ◽  
Author(s):  
Hongwei Yao ◽  
Michelle W. Lee ◽  
Alan J. Waring ◽  
Gerard C. L. Wong ◽  
Mei Hong
Keyword(s):  
Fusion Protein ◽  
Transmembrane Domain ◽  
Magic Angle Spinning ◽  
Structure Characterization ◽  
Cubic Phase ◽  
Magic Angle ◽  
Viral Fusion ◽  
Virus Envelope ◽  
Viral Fusion Protein ◽  
Topological Changes

The C-terminal transmembrane domain (TMD) of viral fusion proteins such as HIV gp41 and influenza hemagglutinin (HA) is traditionally viewed as a passive α-helical anchor of the protein to the virus envelope during its merger with the cell membrane. The conformation, dynamics, and lipid interaction of these fusion protein TMDs have so far eluded high-resolution structure characterization because of their highly hydrophobic nature. Using magic-angle-spinning solid-state NMR spectroscopy, we show that the TMD of the parainfluenza virus 5 (PIV5) fusion protein adopts lipid-dependent conformations and interactions with the membrane and water. In phosphatidylcholine (PC) and phosphatidylglycerol (PG) membranes, the TMD is predominantly α-helical, but in phosphatidylethanolamine (PE) membranes, the TMD changes significantly to the β-strand conformation. Measured order parameters indicate that the strand segments are immobilized and thus oligomerized. 31P NMR spectra and small-angle X-ray scattering (SAXS) data show that this β-strand–rich conformation converts the PE membrane to a bicontinuous cubic phase, which is rich in negative Gaussian curvature that is characteristic of hemifusion intermediates and fusion pores. 1H-31P 2D correlation spectra and 2H spectra show that the PE membrane with or without the TMD is much less hydrated than PC and PG membranes, suggesting that the TMD works with the natural dehydration tendency of PE to facilitate membrane merger. These results suggest a new viral-fusion model in which the TMD actively promotes membrane topological changes during fusion using the β-strand as the fusogenic conformation.

scholarly journals Mutations in the Transmembrane Domain and Cytoplasmic Tail of Hendra Virus Fusion Protein Disrupt Virus-Like-Particle Assembly

2017 ◽  
Vol 91 (14) ◽  
Author(s):  
Nicolás Cifuentes-Muñoz ◽  
Weina Sun ◽  
Greeshma Ray ◽  
Phuong Tieu Schmitt ◽  
Stacy Webb ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Transmembrane Domain ◽  
Cytoplasmic Tail ◽  
Hendra Virus ◽  
Endocytic Trafficking ◽  
Viral Fusion ◽  
Particle Assembly ◽  
Particle Release ◽  
F Protein ◽  
Viral Fusion Protein

ABSTRACT Hendra virus (HeV) is a zoonotic paramyxovirus that causes deadly illness in horses and humans. An intriguing feature of HeV is the utilization of endosomal protease for activation of the viral fusion protein (F). Here we investigated how endosomal F trafficking affects HeV assembly. We found that the HeV matrix (M) and F proteins each induced particle release when they were expressed alone but that their coexpression led to coordinated assembly of virus-like particles (VLPs) that were morphologically and physically distinct from M-only or F-only VLPs. Mutations to the F protein transmembrane domain or cytoplasmic tail that disrupted endocytic trafficking led to failure of F to function with M for VLP assembly. Wild-type F functioned normally for VLP assembly even when its cleavage was prevented with a cathepsin inhibitor, indicating that it is endocytic F trafficking that is important for VLP assembly, not proteolytic F cleavage. Under specific conditions of reduced M expression, we found that M could no longer induce significant VLP release but retained the ability to be incorporated as a passenger into F-driven VLPs, provided that the F protein was competent for endocytic trafficking. The F and M proteins were both found to traffic through Rab11-positive recycling endosomes (REs), suggesting a model in which F and M trafficking pathways converge at REs, enabling these proteins to preassemble before arriving at plasma membrane budding sites. IMPORTANCE Hendra virus and Nipah virus are zoonotic paramyxoviruses that cause lethal infections in humans. Unlike that for most paramyxoviruses, activation of the henipavirus fusion protein occurs in recycling endosomal compartments. In this study, we demonstrate that the unique endocytic trafficking pathway of Hendra virus F protein is required for proper viral assembly and particle release. These results advance our basic understanding of the henipavirus assembly process and provide a novel model for the interplay between glycoprotein trafficking and paramyxovirus assembly.

scholarly journals Membrane Interactions of the Tick-Borne Encephalitis Virus Fusion Protein E at Low pH

2002 ◽  
Vol 76 (8) ◽  
pp. 3784-3790 ◽  
Author(s):  
Karin Stiasny ◽  
Steven L. Allison ◽  
Juliane Schalich ◽  
Franz X. Heinz
Keyword(s):  
Fusion Protein ◽  
Membrane Binding ◽  
Low Ph ◽  
Encephalitis Virus ◽  
Soluble Form ◽  
Viral Fusion ◽  
Sequence Element ◽  
Viral Fusion Protein ◽  
Tick Borne Encephalitis ◽  
Tick Borne Encephalitis Virus

ABSTRACT Membrane fusion of the flavivirus tick-borne encephalitis virus is triggered by the mildly acidic pH of the endosome and is mediated by envelope protein E, a class II viral fusion protein. The low-pH trigger induces an oligomeric rearrangement in which the subunits of the native E homodimers dissociate and the monomeric subunits then reassociate into homotrimers. Here we provide evidence that membrane binding is mediated by the intermediate monomeric form of E, generated by low-pH-induced dissociation of the dimer. Liposome coflotation experiments revealed that association with target membranes occurred only when liposomes were present at the time of acidification, whereas pretreating virions at low pH in the absence of membranes resulted in the loss of their ability to stably attach to liposomes. With the cleavable cross-linker ethylene glycolbis(succinimidylsuccinate), it was shown that a truncated soluble form of the E protein (sE) could bind to membranes only when the dimers were free to dissociate at low pH, and binding could be blocked by a monoclonal antibody that recognizes the fusion peptide, which is at the distal tip of the E monomer but is buried in the native dimer. Surprisingly, analysis of the membrane-associated sE proteins revealed that they had formed trimers. This was unexpected because this protein lacks a sequence element in the C-terminal stem-anchor region, which was shown to be essential for trimerization in the absence of a target membrane. It can therefore be concluded that the formation of a trimeric form of sE is facilitated by membrane binding. Its stability is apparently maintained by contacts between the ectodomains only and is not dependent on sequence elements in the stem-anchor region as previously assumed.  

Zirconium oxide nanoparticles coated on sepiolite by sol–gel process — Their application as a solvent-free catalyst for condensation reactions

2011 ◽  
Vol 89 (3) ◽  
pp. 280-288 ◽  
Author(s):  
Sadok Letaief ◽  
Yun Liu ◽  
Christian Detellier
Keyword(s):  
Clay Mineral ◽  
Zirconium Oxide ◽  
Condensation Reaction ◽  
Sol Gel ◽  
Magic Angle Spinning ◽  
Solvent Free ◽  
Cubic Phase ◽  
Magic Angle ◽  
Solvent Free Conditions ◽  
Sol Gel Process

An inorganic nanocomposite made of zirconia nanoparticles coated on the external surfaces of the fibrous clay mineral sepiolite was prepared by using the sol–gel process under soft conditions using zirconium(IV) propoxide in 1-propanol as the precursor. The resulting materials were characterized by X-ray diffraction (XRD), thermal gravimetric analyses (TGA) and differential thermal analyses (DTA), microporosimetry, 29Si magic-angle spinning (MAS) nuclear magnetic resonance (NMR), and high-resolution transmission electron microscopy (HR-TEM). The organic material from the precursor was fully removed after calcination at 450 °C, concurrently with the crystallization of the cubic phase of zirconium oxide. The coordinated water molecules of sepiolite were fully removed and its symmetrical layered structure was folded after calcination at 650 °C, as observed by XRD and 29Si cross-polarization (CP) / MAS NMR. The arrangement of the nanoparticles of zirconium oxide on the sepiolite external surfaces maintains a fibrous morphology for the nanocomposite. This material was applied as a catalyst for the Knoevenagel condensation reaction of benzaldehyde and malononitrile under solvent-free conditions. Strongly improved yields of reaction, attributed to larger catalytically active surfaces, were obtained compared with either the sepiolite clay mineral or the zirconium oxide tested individually.

scholarly journals Role of Metastability and Acidic pH in Membrane Fusion by Tick-Borne Encephalitis Virus

2001 ◽  
Vol 75 (16) ◽  
pp. 7392-7398 ◽  
Author(s):  
Karin Stiasny ◽  
Steven L. Allison ◽  
Christian W. Mandl ◽  
Franz X. Heinz
Keyword(s):  
Influenza Virus ◽  
Fusion Protein ◽  
Membrane Fusion ◽  
Low Ph ◽  
Class I ◽  
Acidic Ph ◽  
Viral Fusion ◽  
Viral Fusion Protein ◽  
Tick Borne Encephalitis ◽  
Tbe Virus

ABSTRACT The envelope protein E of the flavivirus tick-borne encephalitis (TBE) virus is, like the alphavirus E1 protein, a class II viral fusion protein that differs structurally and probably mechanistically from class I viral fusion proteins. The surface of the native TBE virion is covered by an icosahedrally symmetrical network of E homodimers, which mediate low-pH-induced fusion in endosomes. At the pH of fusion, the E homodimers are irreversibly converted to a homotrimeric form, which we have found by intrinsic fluorescence measurements to be more stable than the native dimers. Thus, the TBE virus E protein is analogous to the prototypical class I fusion protein, the influenza virus hemagglutinin (HA), in that it is initially synthesized in a metastable state that is energetically poised to be converted to the fusogenic state by exposure to low pH. However, in contrast to what has been observed with influenza virus HA, this transition could not be triggered by input of heat energy alone and membrane fusion could be induced only when the virus was exposed to an acidic pH. In a previous study we showed that the dimer-to-trimer transition appears to be a two-step process involving a reversible dissociation of the dimer followed by an irreversible trimerization of the dissociated monomeric subunits. Because the dimer-monomer equilibrium in the first step apparently depends on the protonation state of E, the lack of availability of monomers for the trimerization step at neutral pH could explain why low pH is essential for fusion in spite of the metastability of the native E dimer.

scholarly journals Identification and Characterization of the Putative Fusion Peptide of the Severe Acute Respiratory Syndrome-Associated Coronavirus Spike Protein

2005 ◽  
Vol 79 (11) ◽  
pp. 7195-7206 ◽  
Author(s):  
Bruno Sainz ◽  
Joshua M. Rausch ◽  
William R. Gallaher ◽  
Robert F. Garry ◽  
William C. Wimley
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Fusion Protein ◽  
Protein A ◽  
Fusion Peptide ◽  
Class I ◽  
Viral Fusion ◽  
Fusion Peptides ◽  
Viral Fusion Protein ◽  
Ww Ii ◽  
Heptad Repeats

ABSTRACT Severe acute respiratory syndrome-associated coronavirus (SARS-CoV) is a newly identified member of the family Coronaviridae and poses a serious public health threat. Recent studies indicated that the SARS-CoV viral spike glycoprotein is a class I viral fusion protein. A fusion peptide present at the N-terminal region of class I viral fusion proteins is believed to initiate viral and cell membrane interactions and subsequent fusion. Although the SARS-CoV fusion protein heptad repeats have been well characterized, the fusion peptide has yet to be identified. Based on the conserved features of known viral fusion peptides and using Wimley and White interfacial hydrophobicity plots, we have identified two putative fusion peptides (SARSWW-I and SARSWW-II) at the N terminus of the SARS-CoV S2 subunit. Both peptides are hydrophobic and rich in alanine, glycine, and/or phenylalanine residues and contain a canonical fusion tripeptide along with a central proline residue. Only the SARSWW-I peptide strongly partitioned into the membranes of large unilamellar vesicles (LUV), adopting a β-sheet structure. Likewise, only SARSWW-I induced the fusion of LUV and caused membrane leakage of vesicle contents at peptide/lipid ratios of 1:50 and 1:100, respectively. The activity of this synthetic peptide appeared to be dependent on its amino acid (aa) sequence, as scrambling the peptide rendered it unable to partition into LUV, assume a defined secondary structure, or induce both fusion and leakage of LUV. Based on the activity of SARSWW-I, we propose that the hydrophobic stretch of 19 aa corresponding to residues 770 to 788 is a fusion peptide of the SARS-CoV S2 subunit.

scholarly journals Reevaluating Herpes Simplex Virus Hemifusion

2010 ◽  
Vol 84 (22) ◽  
pp. 11814-11821 ◽  
Author(s):  
Julia O. Jackson ◽  
Richard Longnecker
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Fusion Protein ◽  
Membrane Fusion ◽  
Fusion Proteins ◽  
Fusion Pore ◽  
Viral Fusion ◽  
Viral Fusion Protein ◽  
Viral Fusion Proteins ◽  
Simplex Virus

ABSTRACT Membrane fusion induced by enveloped viruses proceeds through the actions of viral fusion proteins. Once activated, viral fusion proteins undergo large protein conformational changes to execute membrane fusion. Fusion is thought to proceed through a “hemifusion” intermediate in which the outer membrane leaflets of target and viral membranes mix (lipid mixing) prior to fusion pore formation, enlargement, and completion of fusion. Herpes simplex virus type 1 (HSV-1) requires four glycoproteins—glycoprotein D (gD), glycoprotein B (gB), and a heterodimer of glycoprotein H and L (gH/gL)—to accomplish fusion. gD is primarily thought of as a receptor-binding protein and gB as a fusion protein. The role of gH/gL in fusion has remained enigmatic. Despite experimental evidence that gH/gL may be a fusion protein capable of inducing hemifusion in the absence of gB, the recently solved crystal structure of HSV-2 gH/gL has no structural homology to any known viral fusion protein. We found that in our hands, all HSV entry proteins—gD, gB, and gH/gL—were required to observe lipid mixing in both cell-cell- and virus-cell-based hemifusion assays. To verify that our hemifusion assay was capable of detecting hemifusion, we used glycosylphosphatidylinositol (GPI)-linked hemagglutinin (HA), a variant of the influenza virus fusion protein, HA, known to stall the fusion process before productive fusion pores are formed. Additionally, we found that a mutant carrying an insertion within the short gH cytoplasmic tail, 824L gH, is incapable of executing hemifusion despite normal cell surface expression. Collectively, our findings suggest that HSV gH/gL may not function as a fusion protein and that all HSV entry glycoproteins are required for both hemifusion and fusion. The previously described gH 824L mutation blocks gH/gL function prior to HSV-induced lipid mixing.

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