title: Conformation states of gramicidin A along the pathway to the formation of channels in model membranes determined by 2D NMR and circular dichroism spectroscopy.
abstract: Gramicidin A incorporated into SDS (sodium dodecyl sulfate) micelles exists as a right-handed, N-to-N-terminal beta 6.3 helical dimer [Lomize, A. L., Orechov, V. Yu., & Arseniev, A.S. (1992) Bioorg. Khim. 18, 182-189]. In the incorporation procedure to achieve the ion channel state of gramicidin A in SDS micelles, trifluoroethanol (TFE) is used to solubilize the hydrophobic peptide before addition to the aqueous/micelle solution. The conformational transition of gramicidin A to form ion channels in SDS micelles, i.e., in TFE and 10% TFE/water, has been investigated using 2D NMR and CD spectroscopy. In neat TFE, gramicidin A was found to be monomeric and may possibly exist in an equilibrium of rapidly interconverting conformers of at least three different forms believed to be left- and/or right-handed alpha and beta 4.4 helices. It was found that the interconversion between these conformers was slowed down in 55% TFE as evident by the observation of at least three different sets of d alpha N COSY peaks although CD gave a net spectrum similar to that in neat TFE. In 10% TFE gramicidin A spontaneously forms a precipitate. The precipitated species were isolated and solubilized in dioxane where gramicidin conformers undergo very slow interconversion and could be characterized by NMR. At least seven different gramicidin A conformations were found in 10% TFE. Four of thes are the same types of double helices as previously found in ethanol (i.e., a symmetric left-handed parallel beta 5.6 double helix, an unsymmetric left-handed parallel beta 5.6 double helix, a symmetric left-handed antiparallel beta 5.6 double helix, a symmetric right-handed parallel beta 5.6 double helix); the fifth is possibly a symmetric right-handed antiparallel beta 5.6 double helix. There is also evidence for the presence of at least one form of monomeric species. Previous observation on the solvent history dependence in the ease of channel incorporation may be explained by the presence of several different folding pathways to channel formation. To test this proposal, the conformation of gramicidin A in 10% DMSO and 10% methanol was studied. In the former environment, the major form was a random coil with a minor population of double-stranded helices, while in the latter, NMR spectra indicate the presence of the same double-helical conformers as found in neat methanol.
authors.affiliation: Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville 72701.
authors.forename: N
authors.initials: N
authors.lastname: Abdul-Manan
authors.forename: J F
authors.initials: JF
authors.lastname: Hinton
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004146:Dioxanes; D006096:Gramicidin / Q000737:chemistry*; D007473:Ion Channels / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008567:Membranes, Artificial; D000432:Methanol; D008823:Micelles; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012997:Solvents; D014270:Trifluoroethanol; D014867:Water
chemical_list: D004146:Dioxanes; D007473:Ion Channels; D008567:Membranes, Artificial; D008823:Micelles; D012997:Solvents; D014867:Water; D006096:Gramicidin; D014270:Trifluoroethanol; C025223:1,4-dioxane; D000432:Methanol