Line-tension controlled mechanism for influenza fusion - PubMed (original) (raw)

Line-tension controlled mechanism for influenza fusion

Herre Jelger Risselada et al. PLoS One. 2012.

Abstract

Our molecular simulations reveal that wild-type influenza fusion peptides are able to stabilize a highly fusogenic pre-fusion structure, i.e. a peptide bundle formed by four or more trans-membrane arranged fusion peptides. We rationalize that the lipid rim around such bundle has a non-vanishing rim energy (line-tension), which is essential to (i) stabilize the initial contact point between the fusing bilayers, i.e. the stalk, and (ii) drive its subsequent evolution. Such line-tension controlled fusion event does not proceed along the hypothesized standard stalk-hemifusion pathway. In modeled influenza fusion, single point mutations in the influenza fusion peptide either completely inhibit fusion (mutants G1V and W14A) or, intriguingly, specifically arrest fusion at a hemifusion state (mutant G1S). Our simulations demonstrate that, within a line-tension controlled fusion mechanism, these known point mutations either completely inhibit fusion by impairing the peptide's ability to stabilize the required peptide bundle (G1V and W14A) or stabilize a persistent bundle that leads to a kinetically trapped hemifusion state (G1S). In addition, our results further suggest that the recently discovered leaky fusion mutant G13A, which is known to facilitate a pronounced leakage of the target membrane prior to lipid mixing, reduces the membrane integrity by forming a 'super' bundle. Our simulations offer a new interpretation for a number of experimentally observed features of the fusion reaction mediated by the prototypical fusion protein, influenza hemagglutinin, and might bring new insights into mechanisms of other viral fusion reactions.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1

Figure 1. Standard stalk-hemifusion pathway (cross-section, side-view): The initial stalk (I) radially expands (II) forming an H-shaped hemifusion diaphragm (H-HD) after the trans-leaflets (colored yellow) meet (III).

When the H-HD ruptures a fusion pore is formed (IV).

Figure 2

Figure 2. A -shaped hemifusion diaphragm (-HD) which is generated by a stalk that has encircled a membrane pore.

Figure 3

Figure 3. Evolution of a stalk in the absence and presence of a pore.

For sake of clarity the size of the lipid headgroups is exaggerated (solvent is not shown). (A) Two apposed DOPC bilayers. A preformed stalk is not stable (see Fig. 4). (B) A stable ‘hour-glass shaped’ stalk structure formed between a DOPE and DOPC bilayer (4 µs). (C) Elongation of a stalk formed between two DOPE bilayers (4 µs). (I-III) Evolution of a stalk formed between two DOPC bilayers in the vicinity of a pore (stalk-pore complex). Elongation of the stalk, which circumvents the pore, results in the formation of a formula image-shaped hemifusion diaphragm (formula image-HD).

Figure 4

Figure 4. Stalk evolution in response of removing the pore (top view on porated bilayer).

(I,II,III) Sudden removal of the pore before completion of the formula image-HD reverses the stalk elongation process, and the stalk completely disappears (DOPC at 310 K, with formula image water molecules per lipid between the membranes). Hydrophobic lipid tails are colored grey, polar-headgroups (DOPC) tan.

Figure 5

Figure 5. (A) Overlap between the coarse-grained model (backbone red and side-chains yellow) of the wild-type influenza fusion peptide and the NMR structure .

The two helices are joined by a linker region at a slightly bent angle (boomerang-shape). (B) The wild-type influenza fusion peptides (side-chains not shown) aggregate into a stable hexameric bundle. The bundle interior is depleted in solvent (colored blue) and lipid head groups. For sake of clarity, the first backbone residue (Gly1) is colored yellow. (C) Top view of the bundle. The bundle’s interior is mainly composed of the hydrophilic residues Glu11 (colored blue) and Asn12 (colored green) that are located in the kinked region of the peptide and which point toward the central axis of the bundle.

Figure 6

Figure 6. Evolution of the stalk in the presence of the peptide bundle.

For sake of clarity the size of the lipid headgroups is exaggerated (solvent is not shown). (A) The elongated stalk (wild-type peptides) after 0.4 formula images. The bundle has opened up and the stalk and has partly surrounded the formed hole. Notice the readily adopted banana-shape. The stalk forces the peptides to the remaining rim portion. At this stage mixing occurs between both the _cis_-leaflets and the _trans_-leaflet of the target membrane (colored gray), while the _cis_-leaflet of the host cell (colored yellow) does not contribute to lipid mixing. (B) Mutating a single residue in the peptides, Gly1 to Ser1 (colored green), known as the terminal hemifusion mutant G1S , , stabilizes both bundle and stalk but inhibits elongation of the stalk (10 formula images). Consequentially, the fusion reaction becomes trapped. Note that lipid head-groups are excluded from the pore interior and the _trans_-leaflets (colored yellow) are hindered from participating in the lipid mixing.

Figure 7

Figure 7. Point mutations that are known to inhibit fusion destabilize the bundle.

(top) Mutating a single residue, Gly1 to Val1 (colored blue), destabilizes the peptide bundle (mutant G1V). (bottom) Mutating a single residue, Trp14 to Ala14 (colored cyan), rapidly destabilizes the peptide bundle (mutant W14A). Notice the flexible kink that points out of the membrane .

Figure 8

Figure 8. The G1S mutation reverses the stalk elongation process facilitated by the wild-type.

Notice the removal of solvent (colored blue) and lipid head-groups (colored tan) from the membrane interior when the peptide bundle ‘reseals’ itself – the stalk and peptide are competitive lineactants.

Figure 9

Figure 9. Interaction between multiple peptide bundles.

(upper panel) Four wild-type bundles (top-view). The bundles strongly repel each other and maximize their separation distance in the course of the simulation. Eventually one of the bundles vanishes. (middle panel) Four G13A mutant bundles. The bundles are attractive and their coalescence results in a ‘super’ bundle consisting of 10 trans-membrane arranged peptides. (lower panel) Aggregation number of the largest bundle in the course of the simulation (Only the trans-membrane arranged peptide are counted). The brown line shows a separate simulation where the G13A mutation is reversed after 20 formula images (G13A -> wild-type). The wild-type ‘super’ bundle readopts its usual size in the course of the simulation.

Figure 10

Figure 10. Detailed view of the ‘super’ bundle (10 s) formed by the leaky fusion mutant G13A (Top view, cross-section through the bilayer center).

Notice that residue 13 (colored green) directly faces the hydrophobic lipid rim around the bundle. The solvent (colored blue) in the center of the bundle suggests the occurrence of leakage prior to lipid mixing .

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