The Central Proline of an Internal Viral Fusion Peptide Serves Two Important Roles

Delos, Sue E.; Gilbert, Joanna M.; White, Judith M.

doi:10.1128/jvi.74.4.1686-1693.2000

Cited by 68 publications

(79 citation statements)

References 55 publications

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“…Most notably, rather than stabilizing an extended structure that contains the FP, the cysteines in the p10 ectodomain stabilize an 11-residue loop that appears to constitute the actual FP. Unlike the fusion loops of the ␣-retroviruses and Ebola virus, where proline residues may be important for formation of the turn in the loop (47,48), the p10 cystine loop lacks prolines, and loop formation directly relies on formation of the intramolecular disulfide bond. The p10 cystine loop resembles the tips of the fusion loops of the class II and class III enveloped viruses, which exist as open, compact, or flexible loops (49 -51).…”

Section: Discussionmentioning

confidence: 99%

Features of a Spatially Constrained Cystine Loop in the p10 FAST Protein Ectodomain Define a New Class of Viral Fusion Peptides

Barry¹,

Key²,

Haddad³

et al. 2010

Journal of Biological Chemistry

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The reovirus fusion-associated small transmembrane (FAST) proteins are the smallest known viral membrane fusion proteins. With ectodomains of only ϳ20 -40 residues, it is unclear how such diminutive fusion proteins can mediate cell-cell fusion and syncytium formation. Contained within the 40-residue ectodomain of the p10 FAST protein resides an 11-residue sequence of moderately apolar residues, termed the hydrophobic patch (HP). Previous studies indicate the p10 HP shares operational features with the fusion peptide motifs found within the enveloped virus membrane fusion proteins. Using biotinylation assays, we now report that two highly conserved cysteine residues flanking the p10 HP form an essential intramolecular disulfide bond to create a cystine loop. Mutagenic analyses revealed that both formation of the cystine loop and p10 membrane fusion activity are highly sensitive to changes in the size and spatial arrangement of amino acids within the loop. The p10 cystine loop may therefore function as a cystine noose, where fusion peptide activity is dependent on structural constraints within the noose that force solvent exposure of key hydrophobic residues. Moreover, inhibitors of cell surface thioreductase activity indicate that disruption of the disulfide bridge is important for p10-mediated membrane fusion. This is the first example of a viral fusion peptide composed of a small, spatially constrained cystine loop whose function is dependent on altered loop formation, and it suggests the p10 cystine loop represents a new class of viral fusion peptides.Membrane fusion is an essential process involved in an abundance of biological events, including sperm-egg fusion, multinucleated myotube formation, exocytosis, and enveloped virus entry (1). The fusion reaction is catalyzed by specialized membrane fusion proteins designed to alter bilayer structure and bring about membrane merger. The membrane fusion proteins of various enveloped viruses represent some of the best characterized examples of such fusion catalysts (2). Detailed structural and functional analysis of diverse enveloped virus membrane fusion proteins reveals a common pathway of protein-mediated membrane fusion. Triggered exposure of a hydrophobic fusion peptide (FP), 3 normally sequestered within the complex pre-fusion ectodomain structure of these proteins, results in FP insertion into the donor and/or target membranes. Subsequent structural remodeling of these large ectodomains generates a trimeric hairpin structure that draws the two membranes together, driving lipid mixing (also referred to as hemifusion) and pore formation and expansion (3, 4). Despite a wealth of information, the precise mechanisms by which FPs and conformational remodeling of enveloped virus fusion proteins mediate lipid bilayer rearrangement and fusion remain unclear.The reovirus fusion-associated small transmembrane (FAST) proteins are a singular family of viral membrane fusion proteins. Although many nonenveloped viruses encode proteins involved in membrane permeabilization (...

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Section: Discussionmentioning

confidence: 99%

Features of a Spatially Constrained Cystine Loop in the p10 FAST Protein Ectodomain Define a New Class of Viral Fusion Peptides

Barry¹,

Key²,

Haddad³

et al. 2010

Journal of Biological Chemistry

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“…In several viruses with internal fusion peptides, there is a central proline within this sequence. Such a reverse-turn function seems to be related to the efficiency in processing during maturation, as shown by a series of mutations in avian sarcoma/leucosis virus (53). Receptor binding was not much affected by point mutations at this proline site.…”

mentioning

confidence: 95%

“…However, the infectivity showed a bell-shaped dependence viz. the hydrophobicity of the residue, where proline is of a moderate level (53). In SFV the cholesterol dependence for fusion is linked to another proline, Pro-226, also in the E1 glycoprotein.…”

mentioning

confidence: 99%

Prefusion Rearrangements Resulting in Fusion Peptide Exposure in Semliki Forest Virus

Hammar¹,

Markarian²,

Haag³

et al. 2003

Journal of Biological Chemistry

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Semliki Forest virus (SFV), like many enveloped viruses, takes advantage of the low pH in the endosome to convert into a fusion-competent configuration and complete infection by fusion with the endosomal membrane. Unlike influenza virus, carrying an N-terminal fusion peptide, SFV represents a less-well understood fusion principle involving an endosequence fusion peptide. To explore the series of events leading to a fusogenic configuration of the SFV, we exposed the virus to successive acidification, mimicking endosomal conditions, and followed structural rearrangements at probed sensor surfaces. Thus revealed, the initial phase involves a transient appearance of a non-linear neutralizing antibody epitope in the fusion protein, E1. Concurrent with the disappearance of this epitope, a set of masked sequences in proteins E1 and E2 became exposed. When pH reached 6.0 -5.9 the virion transformed into a configuration of enlarged diameter with the fusion peptide optimally exposed. Simultaneously, a partly hidden sequence close to the receptor binding site in E2 became fully uncovered. At this presumably fusogenic stage, maximally 80 fusion peptide-identifying antibody Fab fragments could be bound per virion, i.e. one ligand per three copies of the fusion protein. The phenomena observed are discussed in terms of alphavirus structure and reported functional domains.

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“…Peptides corresponding to the membrane-interacting portions of several viral glycoproteins have been functionally and structurally characterized (reviewed in reference 42). Many of these HRs share a common feature: the presence of a critical proline at or near the center (9,11). This proline likely induces these regions to sit in the membrane as kinked structures.…”

Section: Sequence Analysis Of the Pnrsv Mpmentioning

confidence: 99%

Plant Virus Cell-to-Cell Movement Is Not Dependent on the Transmembrane Disposition of Its Movement Protein

Martínez-Gil

Sánchez‐Navarro

Cruz

et al. 2009

J Virol

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The cell-to-cell transport of plant viruses depends on one or more virus-encoded movement proteins (MPs). Some MPs are integral membrane proteins that interact with the membrane of the endoplasmic reticulum, but a detailed understanding of the interaction between MPs and biological membranes has been lacking. The cell-to-cell movement of the Prunus necrotic ringspot virus (PNRSV) is facilitated by a single MP of the 30K superfamily. Here, using a myriad of biochemical and biophysical approaches, we show that the PNRSV MP contains only one hydrophobic region (HR) that interacts with the membrane interface, as opposed to being a transmembrane protein. We also show that a proline residue located in the middle of the HR constrains the structural conformation of this region at the membrane interface, and its replacement precludes virus movement.Plant viruses encode movement proteins (MPs) that mediate the intra-and intercellular spread of the viral genome via plasmodesmata, membranous channels that traverse the walls of plant cells and enable intercellular transport and communication. There is a range of diversity in the number and type of viral proteins required for viral movement (21). Research on tobacco mosaic virus (TMV) has played a leading role in understanding MP activity (2). The genome of TMV encodes a single 30-kDa multidomain protein, the namesake of the 30K superfamily (7). Viral RNA is associated with the membrane of the endoplasmic reticulum (ER) and microtubules in the presence of this MP (23,30).A large number of plant viruses have 30K MPs, which share common abilities, including binding nucleic acids, localizing and increasing the size exclusion limit of plasmodesmata, and interacting with the ER membrane. A topological model has been proposed in which the TMV MP has two putative transmembrane (TM) helices, both the N and C termini oriented toward the cytoplasm, and a short loop exposed in the ER lumen (4). There is less experimental information for other 30K MPs, but they are likely to have some membrane interaction.Direct experimental evidence of the integration of MPs into the membrane has been obtained only for small hydrophobic MPs that do not belong to the 30K superfamily. There are two TM segments in the p9 protein of carnation mottle virus (41), whereas the p6 protein of beet yellow virus (29) and the p7B protein of melon necrotic spot virus (22) have a single TM segment. In viruses with genomes that include three partially overlapping open reading frames, termed the triple-gene block (TGB), all three TGB proteins are required for movement where the two smaller proteins, TGBp2 and TGBp3, are also TM proteins (24). Furthermore, cross-linking experiments with carnation mottle virus p9 protein demonstrated that its membrane insertion occurs cotranslationally in a signal recognition particle-dependent manner and throughout the cellular membrane integration components, the translocon (33, 34).Prunus necrotic ringspot virus (PNRSV) is a tripartite, positive-strand RNA virus in the genus Ilarvirus ...

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The Central Proline of an Internal Viral Fusion Peptide Serves Two Important Roles

Cited by 68 publications

References 55 publications

Features of a Spatially Constrained Cystine Loop in the p10 FAST Protein Ectodomain Define a New Class of Viral Fusion Peptides

Features of a Spatially Constrained Cystine Loop in the p10 FAST Protein Ectodomain Define a New Class of Viral Fusion Peptides

Prefusion Rearrangements Resulting in Fusion Peptide Exposure in Semliki Forest Virus

Plant Virus Cell-to-Cell Movement Is Not Dependent on the Transmembrane Disposition of Its Movement Protein

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