Topological equilibria of ion channel peptides in oriented lipid bilayers revealed by 15N solid-state NMR spectroscopy.
Ion channel peptides have been prepared by solid-phase peptide synthesis, labeled with 15N at selected sites, and reconstituted into oriented lipid bilayers. The (Leu-Ser-Ser-Leu-Leu-Ser-Leu)3-CONH2 peptide has previously been shown to exhibit well-defined and discrete ionic conductances when investigated by single-channel measurements [Lear, J. D., et al. (1988) Science 240, 1177]. Proton-decoupled 15N solid-state NMR spectroscopy indicates that (Leu-Ser-Ser-Leu-Leu-Ser-Leu)3-CONH2 preferentially aligns parallel to the membrane surface in excellent agreement with its amphipathic helical structure. However, by carefully choosing the conditions of the membrane environment, significant contributions that are indicative of transmembrane alignments become obvious in the 15N chemical shift solid-state NMR spectra. The data thereby provide experimental evidence for an equilibrium between in-plane and transmembrane-oriented helix configurations where the transmembrane and surface-oriented peptide fractions are in slow exchange. Similar topological equilibria are observed when the N-terminus of the LS21 peptide is acetylated. These observations provide experimental support for previous models, suggesting that the channels observed in single-channel conductance measurements are indeed formed by hexameric transmembrane helical bundles. In contrast, the shorter peptide (Leu-Ser-Ser-Leu-Leu-Ser-Leu)2-CONH2 is oriented parallel to the membrane surface under all conditions tested. This peptide exhibits erratic conductance changes when investigated by electrophysiological methods, probably because it is too short to span the lipid bilayer.
Sudheendra
U S
US
Faculté de Chimie, Institut le Bel, Université Louis Pasteur/CNRS FRE2446, 4, rue Blaise Pascal, 67070 Strasbourg, France.
Bechinger
Burkhard
B
D007473:Ion Channels / Q000737:chemistry* / Q000378:metabolism*; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism*; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism*
D007473:Ion Channels; D008051:Lipid Bilayers; D009587:Nitrogen Isotopes; D010455:Peptides