| Abstract
| - The fusion peptide of HIV-1 gp41 is formed by the 16 N-terminal residues of the protein.This 16-amino acid peptide, in common with several other viral fusion peptides, caused a reduction in thebilayer to hexagonal phase transition temperature of dipalmitoleoylphosphatidylethanolamine (TH),suggesting its ability to promote negative curvature in membranes. Surprisingly, an elongated peptidecorresponding to the 33 N-terminal amino acids raised TH, although it was more potent than the 16-aminoacid fusion peptide in inducing lipid mixing with large unilamellar liposomes of 1:1:1 dioleoylphosphatidylethanolamine/dioleoylphosphatidylcholine/cholesterol. The 17-amino acid C-terminal fragment of thepeptide can induce membrane fusion by itself, if it is anchored to a membrane by palmitoylation of theamino terminus, indicating that the additional 17 hydrophilic amino acids contribute to the fusogenicpotency of the peptide. This is not solely a consequence of the palmitoylation, as a random peptide withthe same amino acid composition with a palmitoyl anchor was less potent in promoting membrane fusionand palmitic acid itself had no fusogenic activity. The 16-amino acid N-terminal fusion peptide and thelonger 33-amino acid peptide were labeled with NBD. Fluorescence binding studies indicate that bothpeptides bind to the membrane with similar affinities, indicating that the increased fusogenic activity ofthe longer peptide was not a consequence of a greater extent of membrane partitioning. We also determinedthe secondary structure of the peptides using FTIR spectroscopy. We find that the amino-terminal fusionpeptide is inserted into the membrane as a β-sheet and the 17 C-terminal amino acids lie on the surfaceof the membrane, adopting an α-helical conformation. It was further demonstrated with the use ofrhodamine-labeled peptides that the 33-amino acid peptide self-associated in the membrane while the16-amino acid N-terminal peptide did not. Thus, the 16-amino acid N-terminal fusion peptide of HIVinserts into the membrane and, like other viral fusion peptides, lowers TH. In addition, the 17 consecutiveamino acids enhance the fusogenic activity of the fusion peptide presumably by promoting itsself-association.
|
