15N NMR study of the ionization properties of the influenza virus fusion peptide in zwitterionic phospholipid dispersions.
Influenza virus hemagglutinin (HA)-mediated membrane fusion involves insertion into target membranes of a stretch of amino acids located at the N-terminus of the HA(2) subunit of HA at low pH. The pK(a) of the alpha-amino group of (1)Gly of the fusion peptide was measured using (15)N NMR. The pK(a) of this group was found to be 8.69 in the presence of DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine). The high value of this pK(a) is indicative of stabilization of the protonated form of the amine group through noncovalent interactions. The shift reagent Pr(3+) had large effects on the (15)N resonance from the alpha-amino group of Gly(1) of the fusion peptide in DOPC vesicles, indicating that the terminal amino group was exposed to the bulk solvent, even at low pH. Furthermore, electron paramagnetic resonance studies on the fusion peptide region of spin-labeled derivatives of a larger HA construct are consistent with the N-terminus of this peptide being at the depth of the phosphate headgroups. We conclude that at both neutral and acidic pH, the N-terminal of the fusion peptide is close to the aqueous phase and is protonated. Thus neither a change in the state of ionization nor a significant increase in membrane insertion of this group is associated with increased fusogenicity at low pH.
Zhou
Z
Z
Department of Biochemistry, McMaster University, Hamilton, Ontario L8N 3Z5, Canada.
Macosko
J C
JC
Hughes
D W
DW
Sayer
B G
BG
Hawes
J
J
Epand
R M
RM
D055592:Biophysical Phenomena; D001703:Biophysics; D004578:Electron Spin Resonance Spectroscopy; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D009682:Magnetic Resonance Spectroscopy; D009975:Orthomyxoviridae / Q000737:chemistry*; D010713:Phosphatidylcholines / Q000737:chemistry; D012997:Solvents; D014760:Viral Fusion Proteins / Q000737:chemistry*
D019267:Hemagglutinin Glycoproteins, Influenza Virus; D010713:Phosphatidylcholines; D012997:Solvents; D014760:Viral Fusion Proteins; C017251:1,2-oleoylphosphatidylcholine