Ionophore properties of a synthetic alpha-helical transmembrane fragment of the mitochondrial H+ ATP synthetase of Saccharomyces cerevisiae. Comparison with alamethicin.
A 22-amino acid polypeptide was synthesized to model the central transmembrane segment of subunit 8 of the H+ ATP synthetase of Saccharomyces cerevisiae and to test ionophore properties. Solid-phase synthesis was conducted on benzhydrilamino resin, and purification followed by high pressure liquid chromatography allowed the isolation of the pure product whose NH2 terminal was acetylated and whose molecular weight determined by Fast Atomic Bombardment was the expected 2,666. The infrared spectrum of this peptide in the solid state reveals a fully alpha-helical conformation, whereas in low dielectric constant solvents the alpha-helical content is 60%, as determined by circular dichroism studies. Macroscopic current-voltage curves displayed by different planar lipid bilayers (monomyristoleoyl-glycerol and phosphatidylethanolamine) doped with this peptide suggest a weakly voltage-dependent conductance. Only one conductance level is observed in any given single-channel conductance experiment. However, a series of experiments shows a distribution of conductance states, most often 440 or 3,000 pS, and occasionally 80, 1,200, or 6,500 pS. This behavior contrasts with the usual behavior of alamethicin, chosen as a model of "aggregating-helices" ionophore and whose conductance fluctuates continually between substates, through uptake and release of monomers. Nevertheless, alamethicin too can display, under certain conditions, long-lived and mono-level conductance states similar to those reported here for the newly synthesized peptide. These properties could possibly be explained by the formation of large domains of helical rods with a set of allowed and independent ionic pathways.
D000408:Alamethicin; D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents; D002462:Cell Membrane / Q000201:enzymology; D007473:Ion Channels / Q000502:physiology; D046911:Macromolecular Substances; D008564:Membrane Potentials; D008954:Models, Biological; D010446:Peptide Fragments / Q000302:isolation & purification; D011487:Protein Conformation; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D000900:Anti-Bacterial Agents; D007473:Ion Channels; D046911:Macromolecular Substances; D010446:Peptide Fragments; D000408:Alamethicin; D006180:Proton-Translocating ATPases