Influence of Flanking Residues on Tilt and Rotation Angles of Transmembrane Peptides in Lipid Bilayers. A Solid-State ²H NMR Study

Abstract

To gain insight into the parameters that determine the arrangement of proteins in membranes, 2 H NMR experiments were performed to analyze tilt and rotation angles of membrane-spanning R-helical model peptides upon incorporation in diacylphosphatidylcholine bilayers with varying thickness. The peptides consisted of the sequence acetyl-GW 2 (LA) 8 LW 2 A-NH 2 (WALP23) and analogues thereof, in which the interfacial Trp residues were replaced by Lys (KALP23) and/or the hydrophobic sequence was replaced by Leu (WLP23 and KLP23). The peptides were synthesized with a single deuterium-labeled alanine at four different positions along the hydrophobic segment. For all peptides a small but systematic increase in tilt angle was observed upon decreasing the bilayer thickness. However, significantly larger tilt angles were obtained for the Lys-flanked KALP23 than for the Trp-flanked WALP23, suggesting that interfacial anchoring interactions of Trp may inhibit tilting. Increasing the hydrophobicity resulted in an increase in tilt angle for the Trp-flanked analogue only. For all peptides the maximum tilt angle obtained was remarkably small (less than 12°), suggesting that further tilting is inhibited, most likely due to unfavorable packing of lipids around a tilted helix. The results furthermore showed that the direction of tilt is determined almost exclusively by the flanking residues: Trp-and Lys-flanked peptides were found to have very different rotation angles, which were influenced significantly neither by hydrophobicity of the peptides nor by the extent of hydrophobic mismatch. Finally, very small changes in the side chain angles of the deuterated alanine probes were observed in Trp-flanked peptides, suggesting that these peptides may decrease their hydrophobic length to help them to adapt to thin membranes.Membrane proteins carry out many essential functions in cells. In the specific case of integral proteins, the membrane spanning parts of these molecules are in direct contact with acyl chains and headgroups of the surrounding lipids. Consequently, the lipid environment can modulate structure and dynamics and, hence, activity of membrane proteins (1). For example, changes in hydrophobic thickness of a lipid bilayer can affect the structure of an integral membrane protein by causing a hydrophobic mismatch between the bilayer thickness and the length of the membrane spanning parts of that protein (reviewed in refs 2-4). Understanding the molecular basis of how lipids influence membrane proteins requires precise information on structural properties of transmembrane segments of proteins and knowledge of how these properties are sensitive to protein and lipid composition. It is most suitable to perform such experiments on model membranes of synthetic lipids and transmembrane peptides, since the composition of both the lipids and the peptides can be systematically varied. Model membranes, made of phosphatidylcholine derivatives (PC) 1 and model transmembrane peptides, have already been extensively studied to inves...

Mode of Membrane Interaction and Fusogenic Properties of a de Novo Transmembrane Model Peptide Depend on the Length of the Hydrophobic Core

“…This prediction is in accordance with the FTIR data of this study and with previous studies showing that Ala/ Leu peptides of different lengths are transmembrane helices in membranes (11,16,17,30,35).…”

Variation of the Lateral Mobility of Transmembrane Peptides with Hydrophobic Mismatch

“…The peptide "pinching" behavior shows here its limits, contrary to what was observed for the leucine transmembrane peptide in the lamellar phase made of dodecaneswollen C 12 E 4 bilayers. 22 In addition, the jump in diffusion coefficient values is observed sooner (d π + 7 Å) in PBS buffer (pH 8) than in pure water (d π + 10 Å) for L 12 . This is consistent with the fact that at higher pH, the lysine residues are deprotonated, lowering the energetic cost of translocation inside the membrane.…”

Mesoscopic models of biological membranes

“…The occurrence of lateral inhomogeneities in the membrane may, among others, cause membrane fusion, as suggested by Nielsen et al [208], and it may affect passive permeability to small ions or solutes. There may be mismatch effects which are lipid-mediated, such as the tilting (or even bending) of a whole protein/peptide to adapt to a too thin bilayer [181,[209][210][211][212][213][214][215][216], or even tilting of the individual helices which form a protein. There is indeed some indirect experimental evidence that this is the case for channel proteins [180] and even proteins like Rhodopsin [217], suggesting that a change of the tilt-angle of the individual helices could cause changes in protein activity.…”