Mutational analysis of the putative fusion domain of Ebola virus glycoprotein

Ebola viruses contain a single glycoprotein (GP) spike, which functions as a receptor binding and membrane fusion protein. It contains a highly conser ved hydrophobic region (amino acids 524 to 539) located 24 amino acids down stream of the N terminus of the Ebola virus GP2. subunit. Comparison of thi s region with the structural features of the transmembrane subunit of avian retroviral GPs suggests that the conserved Ebola virus hydrophobic region may, in fact, serve as the fusion peptide. To test this hypothesis directly , we introduced conservative (alanine) and nonconservative (arginine) amino acid substitutions at eight positions in this region of the GP2 molecule. The effects of these mutations were deduced from the ability of the Ebola v irus GP to complement the infectivity of a vesicular stomatitis virus (VSV) lacking the receptor-binding G protein. Some mutations, such as Ile-to-Arg substitutions at positions 532 (I532R), F535R, G536A, and P537R, almost co mpletely abolished the ability of the GP to support VSV infectivity without affecting the transport of GP to the cell surface and its incorporation in to virions or the production of virus particles. Other mutations, such as G 528R, L529A, L529R, I532A, and F535A, reduced the infectivity of the VSV-Eb ola virus pseudotypes by at least one-half. These findings, together with p revious reports of liposome association with a peptide corresponding to pos itions 524 to 539 in the GP molecule, offer compelling support for a fusion peptide role for the conserved hydrophobic region in the Ebola virus GP.