Читать книгу Principles of Virology - Jane Flint, S. Jane Flint - Страница 236

Studies with influenza virus envelope glycoproteins indicate that the initial rate of fusion depends on the surface density of HA, suggesting that clustering of several transmembrane protein trimers is required. The number of envelope trimers required to mediate fusion may vary depending on the virus studied, and is frequently debated. One has to bear in mind that assays measuring fusion rely on the use of artificial membrane-forming lipids in vitro; although they are useful tools to probe the mechanism of fusion, they might not reflect accurately the conditions required for fusion between viral and cellular membranes. Indeed, membrane composition is known to affect the fusion rate.

Fusion proceeds by a hemifusion intermediate where the membrane outer, but not inner, leaflets fuse. Such intermediates can be trapped when the HA membrane-spanning region is replaced by a lipid anchor (Box 5.3). The mechanism by which hemifusion progresses to complete fusion is unknown but does not appear very efficient. It was estimated that only 40% of events causing lipid mixing result in mixing of large aqueous molecules. Initially a small aqueous connection between the two membranes, referred to as a fusion pore, opens abruptly. The nascent fusion pore appears unstable and opens and closes repeatedly, flickers, and can ultimately close or remain open and small or open and dilate. Pore widening occurs either by assembly of several small fusion pores or by expansion of individual small pores to allow mixing of large aqueous molecules. Studies with HA proteins bearing amino acid changes in the fusion peptide demonstrated that this peptide plays an important role in pore widening.

A possible model of how fusion is completed could be provided by a comparison of viral fusion proteins with those found in cellular transport vesicles known as SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors). The pairing of vesicle v-SNAREs to target membrane t-SNAREs drives membrane fusion and delivery of the vesicle cargo to its intracellular or extracellular destination. Each SNARE consists of two domains, a coiled coil and a transmembrane domain. The coiled coils of SNAREs positioned on two different membranes zip up, similar to the hairpin structure identified in viral fusion proteins. This zipping up releases the free energy required to mediate fusion between the two membranes (Fig. 5.17).