title: Interaction of D-amino acid incorporated analogues of pardaxin with membranes.
abstract: The influence of specific L- to D-amino acid substitutions on the interaction of pardaxin, a shark repellent neurotoxin polypeptide, with phospholipid vesicles and human erythrocytes is described. Twelve modified, truncated, or fluorescently labeled [with the fluorophore 7-nitrobenz-2-oxa-1,3-diazole-4-yl (NBD) at their N-terminal amino acid] analogues of pardaxin were synthesized by a solid-phase method. Fluorescence measurements were used to monitor the interaction of the analogues with membranes [Rapaport, D., & Shai, Y. (1991) J. Biol. Chem. 266, 23769-23775]. Upon titration of solutions containing the NBD-labeled peptides with small unilamellar vesicles, the fluorescent emission spectra of all NBD-labeled peptides displayed similar blue-shifts, in addition to enhanced intensities, upon relocation of the probe to the more apolar environment. Binding isotherms were constructed from which surface partition constants, in the range of 10(4) M-1, were derived. The existence of an aggregation process, suggested by the shape of the binding isotherms, could be associated only with those analogues in which the N-helix (residues 1-9) was not perturbed. The alpha-helical content of the analogues was estimated by circular dichroism (CD) spectroscopy, both before and after binding to vesicles at neutral pH. The ability of the peptides to dissipate a diffusion potential and to cause calcein release, as well as to lyse human erythrocytes, served to functionally characterize the peptides. The results support a two alpha-helix model, with a bend at position 13, as best describing pardaxin in its membrane-bound state.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Membrane Research and Biophysics, Weizmann Institute of Science, Rehovot, Israel.
authors.forename: Y
authors.initials: Y
authors.lastname: Pouny
authors.forename: Y
authors.initials: Y
authors.lastname: Shai
mesh_terms: D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D002463:Cell Membrane Permeability / Q000187:drug effects; D002942:Circular Dichroism; D004058:Diffusion; D004910:Erythrocyte Membrane / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D005397:Fish Venoms / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D005452:Fluoresceins / Q000378:metabolism; D005453:Fluorescence; D005456:Fluorescent Dyes; D006801:Humans; D008081:Liposomes; D011392:Proline / Q000378:metabolism; D017433:Protein Structure, Secondary; D012995:Solubility; D013237:Stereoisomerism; D014634:Valinomycin / Q000494:pharmacology
chemical_list: D000596:Amino Acids; D005397:Fish Venoms; D005452:Fluoresceins; D005456:Fluorescent Dyes; D008081:Liposomes; D014634:Valinomycin; C017603:pardaxin; D011392:Proline; C007740:fluorexon