1 research outputs found
Solid-State Nuclear Magnetic Resonance Investigation of the Structural Topology and Lipid Interactions of a Viral Fusion Protein Chimera Containing the Fusion Peptide and Transmembrane Domain
The fusion peptide
(FP) and transmembrane domain (TMD) of viral
fusion proteins play important roles during virus–cell membrane
fusion, by inducing membrane curvature and transient dehydration.
The structure of the water-soluble ectodomain of viral fusion proteins
has been extensively studied crystallographically, but the structures
of the FP and TMD bound to phospholipid membranes are not well understood.
We recently investigated the conformations and lipid interactions
of the separate FP and TMD peptides of parainfluenza virus 5 (PIV5)
fusion protein F using solid-state nuclear magnetic resonance. These
studies provide structural information about the two domains when
they are spatially well separated in the fusion process. To investigate
how these two domains are structured relative to each other in the
postfusion state, when the ectodomain forms a six-helix bundle that
is thought to force the FP and TMD together in the membrane, we have
now expressed and purified a chimera of the FP and TMD, connected
by a Gly-Lys linker, and measured the chemical shifts and interdomain
contacts of the protein in several lipid membranes. The FP–TMD
chimera exhibits α-helical chemical shifts in all the membranes
examined and does not cause strong curvature of lamellar membranes
or membranes with negative spontaneous curvature. These properties
differ qualitatively from those of the separate peptides, indicating
that the FP and TMD interact with each other in the lipid membrane.
However, no <sup>13</sup>C–<sup>13</sup>C cross peaks are observed
in two-dimensional correlation spectra, suggesting that the two helices
are not tightly associated. These results suggest that the ectodomain
six-helix bundle does not propagate into the membrane to the two hydrophobic
termini. However, the loosely associated FP and TMD helices are found
to generate significant negative Gaussian curvature to membranes that
possess spontaneous positive curvature, consistent with the notion
that the FP–TMD assembly may facilitate the transition of the
membrane from hemifusion intermediates to the fusion pore