The Aromatic Domain of the Coronavirus Class I Viral Fusion Protein Induces Membrane Permeabilization: Putative Role during Viral Entry †

Department of Biochemistry, Tulane University, New Orleans, Louisiana, United States
Biochemistry (Impact Factor: 3.02). 02/2005; 44(3):947-58. DOI: 10.1021/bi048515g
Source: PubMed
Coronavirus (CoV) entry is mediated by the viral spike (S) glycoprotein, a class I viral fusion protein. During viral and target cell membrane fusion, the heptad repeat (HR) regions of the S2 subunit assume a trimer-of-hairpins structure, positioning the fusion peptide in close proximity to the C-terminal region of the ectodomain. The formation of this structure appears to drive apposition and subsequent fusion of viral and target cell membranes; however, the exact mechanism is unclear. Here, we characterize an aromatic amino acid rich region within the ectodomain of the S2 subunit that both partitions into lipid membranes and has the capacity to perturb lipid vesicle integrity. Circular dichroism analysis indicated that peptides analogous to the aromatic domains of the severe acute respiratory syndrome (SARS)-CoV, mouse hepatitis virus (MHV) and the human CoV OC43 S2 subunits, did not have a propensity for a defined secondary structure. These peptides strongly partitioned into lipid membranes and induced lipid vesicle permeabilization at peptide/lipid ratios of 1:100 in two independent leakage assays. Thus, partitioning of the peptides into the lipid interface is sufficient to disorganize membrane integrity. Our study of the S2 aromatic domain of three CoVs provides supportive evidence for a functional role of this region. We propose that, when aligned with the fusion peptide and transmembrane domains during membrane apposition, the aromatic domain of the CoV S protein functions to perturb the target cell membrane and provides a continuous track of hydrophobic surface, resulting in lipid-membrane fusion and subsequent viral nucleocapsid entry.

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    • "It is noteworthy that a straight helical structure of a 19-residue long PTM peptide of HIV fusion protein has been determined in DPC micelle solution [76]. The aromatic rich region close to the transmembrane domain is a conserved feature among type I fusion proteins and potentially involved in membrane fusion [56]. Virus–host membrane fusion mechanism is a complex process whereby multiple regions of the fusion protein might be involved in membrane interactions [5,20,22]. "
    [Show abstract] [Hide abstract] ABSTRACT: Severe acute respiratory syndrome-associated coronavirus (SARS-CoV) poses a serious public health hazard. The S2 subunit of the S glycoprotein of SARS-CoV carries out fusion between the virus and the host cells. However, the exact mechanism of the cell fusion process is not well understood. Current model suggests that a conformational transition, upon receptor recognition, of the two heptad core regions of S2 may expose the hydrophobic fusogenic peptide or fusion peptide for membrane insertion. Three regions of the S2 subunit have been proposed to be involved in cell-cell fusion. The N-terminal fusion peptide (FP, residues 770-788), an internal fusion peptide (IFP, residues 873-888) and the pre-transmembrane region (PTM, residues 1185-1202) demonstrated interactions with model lipid membranes and potentially involved in the fusion process. Here, we have determined atomic resolution structures of these three peptides in DPC detergent micelles by solution NMR. FP assumes α-helical conformation with significant distortion at the central Gly residues; enabling a close packing among sidechains of aromatic residues including W, Y and F. The 3-D structure of PMT is characterized by a helix-loop-helix with extensive aromatic interactions within the helices. IFP adopts a rather straight α-helical conformation defined by packing among sidechains of aromatic and aliphatic residues. Paramagnetic spin labeled NMR has demonstrated surface localization of PMT whereas FP and IFP inserted into the micelles. Collectively, data presented in this study will aid in understanding fusion mechanism of SARS-CoV. Copyright © 2014. Published by Elsevier B.V.
    Full-text · Article · Dec 2014 · Biochimica et Biophysica Acta (BBA) - Biomembranes
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    • "Peptides corresponding to this region have also been studied in biochemical assays by other groups [101]. In addition, a third, aromatic region adjacent to the transmembrane domain (the membrane-proximal domain) has been shown to be important in SARS-CoV fusion102103104105. This membrane-proximal domain likely acts in concert with a fusion peptide in the S2 ectodomain to mediate final bilayer fusion once conformational changes have exposed the fusion peptide in the ectodomain. "
    [Show abstract] [Hide abstract] ABSTRACT: Coronaviruses are enveloped positive-stranded RNA viruses that replicate in the cytoplasm. To deliver their nucleocapsid into the host cell, they rely on the fusion of their envelope with the host cell membrane. The spike glycoprotein (S) mediates virus entry and is a primary determinant of cell tropism and pathogenesis. It is classified as a class I fusion protein, and is responsible for binding to the receptor on the host cell as well as mediating the fusion of host and viral membranes-A process driven by major conformational changes of the S protein. This review discusses coronavirus entry mechanisms focusing on the different triggers used by coronaviruses to initiate the conformational change of the S protein: receptor binding, low pH exposure and proteolytic activation. We also highlight commonalities between coronavirus S proteins and other class I viral fusion proteins, as well as distinctive features that confer distinct tropism, pathogenicity and host interspecies transmission characteristics to coronaviruses.
    Full-text · Article · Jun 2012 · Viruses
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    • "The membrane fusion process is energetically favorable and essentially irreversible, but has a considerable kinetic energy barrier [1]. These proteins allow rapid membrane fusion by drawing the opposing membranes together and either stabilizing or providing the activation energy to surmount the transition state [1,2]. In this way, they behave in many aspects like a fusion catalyst. "
    [Show abstract] [Hide abstract] ABSTRACT: Author Summary Virus surface proteins mediate interactions with target cells during the initial events in the process of infection. Inhibiting these proteins is therefore a major target for the development of antiviral drugs. However, there are a very large number of different viruses, each with their own distinct surface proteins and, with just a few exceptions, it is not clear how to build novel molecules to inhibit them. Here we applied a computational binding optimization strategy to an atomic resolution structure of dengue virus serotype 2 envelope protein to generate peptide sequences that should interact strongly with this protein. We picked dengue virus as a target because it is the causative agent for the most important mosquito transmitted viral disease. Out of a small number of candidates designed and tested, we identified two different highly inhibitory peptides. To verify our results, we showed that these peptides block virus:cell binding, interfere with a step during viral entry, alter the surface structure of dengue viral particles, and that they interact directly with dengue virus envelope protein. We expect that our approach may be generally applicable to other viral surface proteins where a high resolution structure is available.
    Full-text · Article · Jun 2010 · PLoS Neglected Tropical Diseases
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