Use of Transmembrane Peptides to Investigate Arginine Interactions with Lipid Bilayers

Abstract

With a pK > 12, the guanidinium side chain of arginine (Arg; R) is positively charged over a very wide pH range. There has been much recent discussion concerning the energetics of Arg inserting into a lipid bilayer, or Arg crossing a lipid bilayer. The topic holds significant intrinsic intellectual interest and also is important for understanding the gating mechanism of voltage-dependent transmembrane channels. We address this problem by direct experimental observation using a designed transmembrane peptide that has interfacial tryptophan (Trp; W) anchors. Within membrane-spanning, alpha-helical GWALP23, acetyl-GGALW5LALALAL12ALALALW19LAGA-amide (Vostrikov, et al. 2008. J Am Chem Soc 130, 12584), we have substituted either R12 or R14 near the putative helix midpoint. Models suggest that the W5 and W19 side chains project from essentially the same side of the GWALP23 alpha-helix, with R14 projecting from the opposite face and R12 from the same face as the W5 and W19 anchors. The R12 side chain in effect is situated between the W anchors. Based upon solid-state NMR spectra from oriented lipid/peptide samples, specifically the deuterium quadrupolar splittings from several 2H-labeled alanines in each Arg-containing peptide, the properties of these sequence isomers depend heavily upon the location of the Arg. We find that GWALP23-R14 adopts a transmembrane orientation with a tilt of about 17° in DOPC (compared to a tilt of about 6° for GWALP23 itself). By contrast, GWALP23-R12 seems to assume several different orientations with respect to a hydrated bilayer of DOPC; one or more of these orientations may represent surface-bound peptide.