Characterization of gramicidin A in an inverted micellar environment. A combined high-performance liquid chromatographic and spectroscopic study.
We have investigated the conformational adaptability of gramicidin A incorporated into reverse micelles of sodium bis(2-ethylhexyl)sulfosuccinate (AOT)/isooctane/water, a so far unexplored "host" membrane-mimetic model system for this peptide. A high-performance liquid chromatographic strategy previously developed for the study of gramicidin in phospholipid vesicles and normal micelles [Bañó et al. (1989) FEBS Lett. 250, 67; Bañó et al. (1991) Biochemistry 30, 886] has been successfully extended to this system. The method has permitted the separation of peptide conformational species, namely, double-stranded dimers and monomers, and an accurate quantitation of their proportion in the inverted micellar environment. It has been demonstrated that, once inserted in the micelle, the double-stranded dimers undergo a dissociation process toward a thermodynamically stable monomeric configuration, whose monomerization rate constant (k1) is dependent in a bell-shaped manner on the water:surfactant mole ratio, w0. A tight correlation between k1 and the double-stranded dimer backbone conformation has been found from the comparison of chromatographic and circular dichroism data. In addition, fluorescence experiments indicate that the peptide tryptophans are in a rather nonpolar environment, with a restricted accessibility to water-soluble quenchers such as acrylamide.
Salom
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Departament de Bioquímica i Biologia Molecular, Universitat de València, Spain.
Abad
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Braco
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D000595:Amino Acid Sequence; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004143:Dioctyl Sulfosuccinic Acid; D005454:Fluorescence Polarization; D006096:Gramicidin / Q000737:chemistry*; D046911:Macromolecular Substances; D008823:Micelles; D008969:Molecular Sequence Data; D011487:Protein Conformation; D013050:Spectrometry, Fluorescence; D013057:Spectrum Analysis
D046911:Macromolecular Substances; D008823:Micelles; D004143:Dioctyl Sulfosuccinic Acid; D006096:Gramicidin