Half-Anchored WALP Peptides: Effect Of Anchor Position On Peptide Orientation

Abstract

Peptides of the “WALP” family, acetyl-GWW(LA)nLWWA-[ethanol]amide, provide an opportune model for investigating protein/lipid interactions. Because the motional behavior of the N- and C-terminal tryptophan (W) residues is different (van der Wel [2007] Biochemistry, 46:7514), it is of interest to investigate how the positions of the anchoring tryptophans will influence the average peptide orientation. To address this question, we synthesized “half-anchored” WALP peptides having only one pair of anchoring tryptophans at either the amino or carboxy terminus. These peptides are acetyl-GGWW(LA)8-ethanolamide and acetyl-(AL)8WWG-ethanolamide, which we designate as “N-anchored” and “C-anchored”, respectively. The hydrophobic lengths of these peptides are similar to that of WALP23, but unlike WALP23 they are anchored to the lipid bilayer membrane on only one side. We find that the half-anchored WALP peptides incorporate into lipid bilayers and assume defined orientations. Unlike shorter half-anchored analogs that contain only three or four Leu-Ala pairs, these longer peptides with eight Leu-Ala pairs show no signs of aggregation and therefore allow further investigation of the peptide/lipid interactions. Circular dichroism spectra indicate that the N-anchored and C-anchored peptides retain alpha-helical conformations. By incorporating deuterated alanines into the sequences, we were able to determine the average orientation of these peptides within mechanically aligned lipid bilayers using solid-state deuterium NMR. The bilayer length-dependent tilt of these half-anchored peptides, in DOPC, DMPC and DLPC lipid bilayer membranes, appears to be somewhat less than for WALP23. The observed average tilts range between about 1 and 6 degrees from the bilayer normal for the N-anchored and C-anchored peptides, compared to 4-8 degrees for WALP23. The intrinsically small tilt values and single anchoring region suggest that anchor residue interactions with lipid head groups may be important for the magnitude of the peptide tilt.