Helical distortion in tryptophan- and lysine-anchored membrane-spanning α-helices as a function of hydrophobic mismatch: A solid-state deuterium NMR investigation using the geometric analysis of labeled alanines method

Abstract

We used solid-state deuterium NMR spectroscopy and geometric analysis of labeled alanines to investigate the structure and orientation of a designed synthetic hydrophobic, membrane-spanning a-helical peptide that is anchored within phosphatidylcholine (PC) bilayers using both Trp and Lys side chains near the membrane/water interface. The 23-amino-acid peptide consists of an alternating Leu/Ala core sequence that is expected to be α-helical, flanked by aromatic and then cationic anchors at both ends of the peptide: acetyl-GKALW(LA)6LWLAKA-amide (KWALP23). The geometric analysis of labeled alanines method was elaborated to permit the incorporation and assignment of multiple alanine labels within a single synthetic peptide. Peptides were incorporated into oriented bilayers of dilauroyl-(di-C12:0-), dimyristoyl- (di-C14:0-), or dioleoyl- (di-C18:1c-) PC. In the C12:0 and C14:0 lipids, the 2H-NMR quadrupolar splittings for the set of six core alanines could not be fit to a canonical undistorted a-helix. Rather, we found that a model containing a helical distortion, such as a localized discontinuity or "kink" near the peptide and bilayer center, could fit the data for KWALP23 in these shorter lipids. The suggestion of helix distortion was confirmed by 2H-NMR spectra for KWALP23 in which Leu8 was changed to deuterated Ala8. Further analysis involving reexamination of earlier data led to a similar conclusion that acetyl-GWW(LA) 8LWWA-amide (WALP23) is distorted in dilauroyl-PC, allowing significant improvement in the fitting of the 2H-NMR results. In contrast, WALP23 and KWALP23 are well represented as undistorted α-helices in dioleoyl-PC, suggesting that the distortion could be a response to hydrophobic mismatch between peptide and lipids. © 2008 by the Biophysical Society.