A Study of Carbobenzoxy-D-phenylalanine-L-phenylalanine-glycine, an Inhibitor of Membrane Fusion, in Phospholipid Bilayers with Multinuclear Magnetic Resonance

Document Type

Article

Language

eng

Publication Date

5-4-1995

Publisher

Elsevier

Source Publication

Biochimica et Biophysica Acta: Biomembranes

Source ISSN

0005-2736

Abstract

The anti-viral and membrane fusion inhibitor, carbobenzoxy-d-phenylalanine-l-phenylalanine-glycine (ZfFG), was studied in phospholipid bilayers, where earlier studies had indicated this peptide functioned. Multinuclear magnetic resonance (NMR) studies were performed with isotopically labeled peptide. A peptide labeled in the glycine carboxyl with 13C was synthesized, and the isotropic13C-NMR chemical shift of that carbon was measured as a function of pH. A pKa of 3.6 for the carboxyl was determined from the peptide bound to a phosphatidylcholine bilayer. ZfFG inhibits the formation by sonication of highly curved, small unilamellar vesicles. Experiments as a function of pH revealed that this ability of ZfFG was governed by a pKa of 3.7. Therefore the protonation state of the carboxyl of ZfFG appeared to regulate the effectiveness of this anti-viral peptide at destabilizing highly curved phospholipid assemblies. Such destabilization had previously been discovered to be related to the mechanism of the anti-fusion and anti-viral activity of this peptide. The location of the carboxyl of ZfFG in the membrane was probed with paramagnetic relaxation enhancement of the13C spin lattice relaxation of the carboxyl carbon in the glycine of ZfFG (enriched in13C). Results suggested that this carboxyl is at or above the surface of the phospholipid bilayer. The dynamics of the molecule in the membrane were examined with2H-NMR studies of ZfFG, deuterated in the α-carbon protons of the glycine. When ZfFG was bound to membranes of phosphatidylcholine, a sharp2H-NMR spectral component was observed, consistent with a disordering of the glycine methylene segment of the peptide. When ZfFG was bound to N-methyl dioleoylphosphatidylethanolamine (N-methyl DOPE) bilayers at temperatures below 30° C, a large quadrupole splitting was observed. These results suggest that ZfFG likely inhibits membrane fusion from the surface of the lipid bilayer, but not by forming a tight, stoichiometric complex with the phospholipids.

Comments

Biochimica et Biophysica Acta: Biomembranes, Vol. 1235, No. 2 (May 4, 1995): 213-220. DOI.

Andrew R. Dentino was affiliated with the State University of New York at Buffalo at the time of publication.

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