Charged or Aromatic Anchor Residue Dependence of Transmembrane Peptide Tilt

Abstract

The membrane-spanning segments of integral membrane proteins often are flanked by aromatic or charged amino acid residues, which may "anchor" the transmembrane orientation. Single spanning transmembrane peptides such as those of the WALP family, acetyl-GWW(LA) n LWWA-amide, furthermore adopt a moderate average tilt within lipid bilayer membranes. To understand the anchor residue dependence of the tilt, we introduce Leu-Ala "spacers" between paired anchors and in some cases replace the outer tryptophans. The resulting peptides, acetyl-GX 2 ALW(LA) 6 LWLAX 22 A-amide, have Trp, Lys, Arg, or Gly in the two X positions. The apparent average orientations of the core helical sequences were determined in oriented phosphatidylcholine bilayer membranes of varying thickness using solid-state 2 H NMR spectroscopy. When X is Lys, Arg, or Gly, the direction of the tilt is essentially constant in different lipids and presumably is dictated by the tryptophans (Trp 5 and Trp 19 ) that flank the inner helical core. The Leu-Ala spacers are no longer helical. The magnitude of the apparent helix tilt furthermore scales nicely with the bilayer thickness except when X is Trp. When X is Trp, the direction of tilt is less well defined in each phosphatidylcholine bilayer and varies up to 70°among 1,2-dioleoyl-sn-glycero-3-phosphocholine, 1,2-dimyristoyl-sn-glycero-3-phosphocholine, and 1,2-dilauroylsn-glycero-3-phosphocholine bilayer membranes. Indeed, the X ‫؍‬ Trp case parallels earlier observations in which WALP family peptides having multiple Trp anchors show little dependence of the apparent tilt magnitude on bilayer thickness. The results shed new light on the interactions of arginine, lysine, tryptophan, and even glycine at lipid bilayer membrane interfaces.The lipid bilayer environment has a profound influence on the properties of peptides and proteins found within it. It is significant that many membrane-spanning proteins have bands of aromatic and/or positively charged residues at the membrane interface, which could serve as anchors for the protein orientation and promote favorable protein-lipid interactions. This anchoring is a widespread characteristic that is observed for a variety of proteins having both ␣ and ␤ transmembrane folds (1-5). Furthermore, polar amino acids influence the topology of membrane proteins as the direction of insertion is driven by the asymmetric positioning of basic residues (Lys and Arg), giving rise to the "positive inside" rule for helical membrane proteins (6). Little is known, nevertheless, about the contributions of these residues in defining the orientations of the transmembrane segments within lipid bilayers (7).Due to the inherent complexity of membrane proteins in the native biological membrane environment, model systems provide meaningful ways to address specific questions about protein/lipid interactions. In particular, model peptides of the WALP family, having the general sequence acetyl-GWW-(LA) n LWWA- [ethanol]amide, have yielded valuable information about peptide orientations, dynam...

Comparative analysis of the orientation of transmembrane peptides using solid-state 2H- and 15N-NMR: mobility matters

“…1c). The low mobility of GWALP23 had been noted before, namely in a comparison with WALP23 using fitted values of S zz in a GALA analysis (Vostrikov et al 2010). It was attributed to the presence of a single Trp residue at each end of the GWALP23 helix, as opposed to two consecutive Trp side-chains at each end in WALP23 (which is also the case of WLP23).…”

Buried lysine, but not arginine, titrates and alters transmembrane helix tilt

“…S1 and Table S1), we analyzed the helix tilt for each of the transmembrane peptides (21,23). (Although excessive peptide dynamics does complicate the determination of helix tilt for some peptides (24)(25)(26), the problem is mitigated and the dynamics become tractable when only two aromatic residues are present (3,21) and additionally when a polar residue is present (17,21). We therefore estimated the rather minimal dynamics of the K12 and K14 peptides using a principal order parameter (21).…”

“…GWALP23 | lysine titration | solid-state deuterium NMR T he basic residues lysine (Lys) and arginine (Arg) in membrane proteins are enriched at the interfacial region and are believed to assist in anchoring the transmembrane orientations (1)(2)(3). In the hydrophobic core of transmembrane proteins, these polar amino acids are significantly depleted because of the energetic cost of burying them in the nonpolar lipid environment.…”

Osteopetrosis Mutation R444L Causes Endoplasmic Reticulum Retention and Misprocessing of Vacuolar H+-ATPase a3 Subunit

“…Asymmetric distribution of basic amino acids, with respect to the lipid bilayer, is a well known factor influencing membrane topology, as has been generalized in the "positive inside rule" of von Heijne (47), and may assist TM membrane insertion (48). Furthermore, Vostrikov et al (49) have shown that Arg, Gly, and aromatic residues near the ends of membrane-spanning ␣-helices may be determinants of helical tilt within the membrane bilayer. Arginine is the most effective, determining tilt direction and inducing tilt angles as high as 24°in synthetic peptides incorporated into artificial membrane systems.…”