Effect of Membrane Curvature-Modifying Lipids on Membrane Fusion by Tick-Borne Encephalitis Virus

Abstract

Enveloped viruses enter cells by fusion of their own membrane with a cellular membrane. Incorporation of inverted-cone-shaped lipids such as lysophosphatidylcholine (LPC) into the outer leaflet of target membranes has been shown previously to impair fusion mediated by class I viral fusion proteins, e.g., the influenza virus hemagglutinin. It has been suggested that these results provide evidence for the stalk-pore model of fusion, which involves a hemifusion intermediate (stalk) with highly bent outer membrane leaflets. Here, we investigated the effect of inverted-cone-shaped LPCs and the cone-shaped oleic acid (OA) on the membrane fusion activity of a virus with a class II fusion protein, the flavivirus tick-borne encephalitis virus (TBEV). This study included an analysis of lipid mixing, as well as of the steps preceding or accompanying fusion, i.e., binding to the target membrane and lipid-induced conformational changes in the fusion protein E. We show that the presence of LPC in the outer leaflet of target liposomes strongly inhibited TBEV-mediated fusion, whereas OA caused a very slight enhancement, consistent with a fusion mechanism involving a lipid stalk. However, LPC also impaired the low-pH-induced binding of a soluble form of the E protein to liposomes and its conversion into a trimeric postfusion structure that requires membrane binding at low pH. Because inhibition is already observed before the lipid-mixing step, it cannot be determined whether impairment of stalk formation is a contributing factor in the inhibition of fusion by LPC. These data emphasize, however, the importance of the composition of the target membrane in its interactions with the fusion peptide that are crucial for the initiation of fusion.Fusion of viral membranes with cellular membranes is a key step in the entry of enveloped viruses into cells. It requires energy to overcome repulsive forces between the opposing membranes and to locally disrupt the original lipid bilayer structures. This energy is provided by structural changes and protein-protein, as well as protein-lipid, interactions of specific viral envelope proteins, designated viral fusion proteins. In infectious virions these proteins exist in a metastable state and respond to a specific fusion trigger (receptor binding or acidic pH) by conformational changes that lead to the exposure of the fusion peptide and the formation of an energetically more stable postfusion structure (23,25,49). So far, two structurally different classes of viral fusion proteins have been identified (30). Class I is represented by the spike-like envelope proteins found in orthomyxo-, paramyxo-, retro-, filo-, and coronaviruses. They are characterized by amino-terminal or amino-proximal fusion peptides, as well as the formation of a hairpin-like postfusion structure with a central trimeric coiled coil. In this postfusion form the membraneinserted N and C termini are located at the same end of a very stable protein rod (11, 51). The class II fusion proteins of flaviviruses and alphaviruses are c...

Crystal Structure of the West Nile Virus Envelope Glycoprotein

“…Roussel (35). Although generally similar, the pH of this transition varies among flaviviruses and is dependent on the lipid composition of the target membrane (40,41). For WNV, the pH at which this change occurs has a peak around pH 6.0 (8,14,21), the same as the pK a of histidine (6.0).…”

5-(Perylen-3-yl)Ethynyl-arabino-Uridine (aUY11), an Arabino-Based Rigid Amphipathic Fusion Inhibitor, Targets Virion Envelope Lipids To Inhibit Fusion of Influenza Virus, Hepatitis C Virus, and Other Enveloped Viruses

“…Enrichment of lipids with polar headgroups of larger diameter than their hydrophobic tails in the outer leaflet favors positive curvature, increasing the activation energy required to form the hemifusion stalk and thereby inhibiting viral fusion (6,36). Accordingly, addition of exogenous lipids of the appropriate shape and polarity (such as LPC) prevents the fusion of many enveloped viruses (8)(9)(10)(37)(38)(39). Phospholipids, however, are not likely to be pharmacologically useful.…”

The Rigid Amphipathic Fusion Inhibitor dUY11 Acts through Photosensitization of Viruses

“…However, the authors present evidence that these RAFIs inhibit virus-cell fusion by stabilizing the positive curvature of membranes by virtue of their molecular geometry: RAFIs are inverted-cone-shaped molecules with a large hydrophilic head group attached to a rigid and planar hydrophobic moiety that inserts into the membrane. Thus, RAFIs are thought to act like similarly shaped lysophospholipids, which are known to impair the critical positive-to-negative membrane transitions that occur during virus-cell fusion (2,7). Nevertheless, we noted that the rigid and planar hydrophobic moiety present in all effective RAFIs is a perylene group (Fig.…”

“…ecently, a few broad-spectrum antivirals have been described that target enveloped virus entry (1)(2)(3)(4)(5)(6)(7)(8)(9)(10). Antivirals that can target the biophysical process of virus-cell membrane fusion itself, rather than any particular viral protein, not only have the potential to be truly broad spectrum (against any enveloped virus) but also will likely have a high barrier to resistance (11)(12)(13)(14).…”