Comparison of Interfacial Tyrosine, Tryptophan and Phenylalanine Residues as Determinants of Orientation and Dynamics of Transmembrane Peptides

Abstract

Aromatic amino acids often flank the transmembrane alpha helices of integral membrane proteins. By favoring locations within the membrane-water interface of the lipid bilayer, the aromatic residues Trp, Tyr, and Phe may serve as anchors to help stabilize a transmembrane orientation. In this work, we compare the influence of interfacial Trp, Tyr or Phe residues upon the properties of helical transmembrane peptides. For such comparisons, it is critical to start with no more than one interfacial aromatic residue near each end of a transmembrane helix, for example that of GWALP23 (acetyl-GGALW5(LA)6LW19LAGA-[ethanol]amide; see J. Biol. Chem. 285, 31723). To this end, we have employed 2H-labeled alanines and solid-state NMR spectroscopy to investigate the consequences of replacing W5 in GWALP23 with Tyr or Phe residues at the same or proximate locations. We find that GWALP23 peptides having Y5, F5, or W5 exhibit essentially the same average tilt in bilayer membranes of DLPC, DMPC, and DOPC. When double Tyr anchors are present, in Y4,5GWALP23, the NMR observables are more subject to dynamic averaging and at the same time less responsive to the bilayer thickness than when a single Tyr or Phe residue occupies position 4 or 5. Interestingly, Y4GWALP23 and Y5GWALP23 show similar low levels of dynamic averaging and have a difference of about 30°-40° in the preferred helix azimuthal rotation angle in each lipid. Decreased dynamics are observed when ring hydrogen bonding is removed as in the case of F4,5GWALP23. We conclude that, in the absence of other functional groups, interfacial aromatic residues determine the preferred orientations and dynamics of membrane-spanning peptides.