Design and characterization of anchoring amphiphilic peptides and their interactions with lipid vesicles.
TL;DR: AALLLAAAAAAAAAAAAAAAAAAAWKKKKKK is identified as the most suitable anchor for the immobilization of lipid vesicles and the secondary structure and membrane affinity of the peptides as well as the effect of the different peptides on the membrane permeability.
Abstract: In an effort to develop a polymer/peptide assembly for the immobilization of lipid vesicles, we have made and characterized four water-soluble amphiphilic peptides designed to associate spontaneously and strongly with lipid vesicles without causing significant leakage from anchored vesicles. These peptides have a primary amphiphilic structure with the following sequences: AAAAAAAAAAAAWKKKKKK, AALLLAAAAAAAAAAAAAAAAAAAWKKKKKK, and KKAALLLAAAAAAAAAAAAAAAAAAAWKKKKKK and its reversed homologue KKKKKKWAAAAA AAAAAAAAAAAAAALLLAAKK. Two of the four peptides have their hydrophobic segments capped at both termini with basic residues to stabilize the transmembrane orientation and to increase the affinity for negatively charged vesicles. We have studied the secondary structure and the membrane affinity of the peptides as well as the effect of the different peptides on the membrane permeability. The influence of the hydrophobic length and the role of lysine residues were clearly established. First, a hydrophobic segment of 24 amino acids, corresponding approximately to the thickness of a lipid bilayer, improves considerably the affinity to zwitterionic lipids compared to the shorter one of 12 amino acids. The shorter peptide has a low membrane affinity since it may not be long enough to adopt a stable conformation. Second, the presence of lysine residues is essential since the binding is dominated by electrostatic interactions, as illustrated by the enhanced binding with anionic lipids. The charges at both ends, however, prevent the peptide from inserting spontaneously in the bilayer since it would involve the translocation of a charged end through the apolar core of the bilayer. The direction of the amino acid sequence of the peptide has no significant influence on its behavior. None of these peptides perturbs membrane permeability even at an incubation lipid to peptide molar ratio of 0.5. Among the four peptides, AALLLAAAAAAAAAAAAAAAAAAAWKKKKKK is identified as the most suitable anchor for the immobilization of lipid vesicles.
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TL;DR: It is shown for phosphatidylcholine bilayers and for other model membranes that these peptides adapt a transmembrane topology without extensive peptide or lipid adaptations under conditions of hydrophobic matching, but that significant rearrangements can result from hydrophilic mismatch.
Abstract: Biological membranes are characterized by a heterogeneous composition, which is not only manifested in the wide variety of their components, but also in aspects like the lateral organization, topology, and conformation of proteins and lipids. In bringing about the correct membrane structure, protein-lipid interactions can be expected to play a prominent role. The extent of hydrophobic matching between transmembrane protein segments and lipids potentially constitutes a versatile director of membrane organization, because a tendency to avoid hydrophobic mismatch could result in compensating adaptations such as tilt of the transmembrane segment or segregation into distinct domains. Also, interfacial interactions between lipid headgroups and the aromatic and charged residues that typically flank transmembrane domains may act as an organizing element. In this review, we discuss the numerous model studies that have systematically explored the influence of hydrophobic matching and interfacial anchoring on membrane structure. Designed peptides consisting of a polyleucine or polyleucine/alanine hydrophobic stretch, which is flanked on both sides by tryptophan or lysine residues, reflect the general layout of transmembrane protein segments. It is shown for phosphatidylcholine bilayers and for other model membranes that these peptides adapt a transmembrane topology without extensive peptide or lipid adaptations under conditions of hydrophobic matching, but that significant rearrangements can result from hydrophobic mismatch. Moreover, these effects depend on the nature of the flanking residues, implying a modulation of the mismatch response by interfacial interactions of the flanking residues. The implications of these model studies for the organization of biomembranes are discussed in the context of recent experiments with more complex systems.
296 citations
Cites background from "Design and characterization of anch..."
...face association with a PC or PC/PG bilayer [39,40]....
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28 Jun 2004
TL;DR: In this article, the authors propose a 2.2.2-approximation algorithm for the 2.1.1-GHz model: 2.3.1/2.
Abstract: 2.
62 citations
TL;DR: The data indicate hydrophobicity values of alanine close to zero when studied in the context of helical polypeptides (> or =24 residues) and phospholipid bilayers and the results are discussed in thecontext of a model where polyalanines adopt a variety of configurations, which are interconnected by multiple equilibria.
54 citations
Cites background from "Design and characterization of anch..."
...A wide variety of polyalanine as well as leucine-alanine peptides have been shown to adopt predominantly helical conformations in membrane environments (Oliver and Deamer, 1994; Zhang et al., 1995; Liu and Deber, 1998; Percot et al., 1999; Harzer and Bechinger, 2000)....
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TL;DR: In the equilibrium states of the peptide/membrane complexes, achieved in time ranges of 50-100 ns, the two peptides behave as expected from experimental and theoretical studies, showing that the spontaneous assembly of peptides and lipids is an unbiased and reliable strategy to produce and study models of equilibrated peptides/lipid complexes of unknown membrane-binding mode and topology.
52 citations
TL;DR: Investigating the influence of the QAC head group and acyl chain length on their permeability-perturbing power and on their affinity for lipidic membranes found that the insertion of QACs inside lipidic bilayers is driven by hydrophobic interactions.
51 citations
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TL;DR: It is found that the interactions of four tryptophan analogues with phosphatidylcholine membranes find that the analogues reside in the vicinity of the glycerol group where they all cause similar modest changes in acyl chain organization and that hydrocarbon penetration was not increased by reduction of hydrogen bonding or electric dipole interaction ability.
Abstract: One of the ubiquitous features of membrane proteins is the preference of tryptophan and tyrosine residues for membrane surfaces that presumably arises from enhanced stability due to distinct interfacial interactions. The physical basis for this preference is widely believed to arise from amphipathic interactions related to imino group hydrogen bonding and/or dipole interactions. We have examined these and other possibilities for tryptophan's interfacial preference by using 1H magic angle spinning (MAS) chemical shift measurements, two-dimensional (2D) nuclear Overhauser effect spectroscopy (2D-NOESY) 1H MAS NMR, and solid state 2H NMR to study the interactions of four tryptophan analogues with phosphatidylcholine membranes. We find that the analogues reside in the vicinity of the glycerol group where they all cause similar modest changes in acyl chain organization and that hydrocarbon penetration was not increased by reduction of hydrogen bonding or electric dipole interaction ability. These observations ...
905 citations
TL;DR: Four alanine-based peptides were designed, synthesized, and tested by circular dichroism for alpha-helix formation in H2O to test for helix stabilization by (Glu-, Lys+) ion pairs or salt bridges (H-bonded ion pairs).
Abstract: Four alanine-based peptides were designed, synthesized, and tested by circular dichroism for alpha-helix formation in H2O. Each peptide has three glutamic/lysine residue pairs, is 16 or 17 amino acids long, and has blocked alpha-NH2 and alpha-COOH groups. In one set of peptides ("i+4"), the glutamic and lysine residues are spaced 4 residues or 1 residue apart. In the other set ("i+3"), the spacing is 3 or 2 residues. Within each of these sets, a pair of peptides was made in which the positions of the glutamic and lysine residues are reversed [Glu, Lys (E,K) vs. Lys, Glu (K,E)] in order to assess the interaction of the charged side chains with the helix dipole. Since the amino acid compositions of these peptides differ at most by a single alanine residue, differences in helicity are caused chiefly by the spacing and positions of the charged residues. The basic aim of this study was to test for helix stabilization by (Glu-, Lys+) ion pairs or salt bridges (H-bonded ion pairs). The results are as follows. (i) All four peptides show significant helix formation, and the stability of the alpha-helix does not depend on peptide concentration in the range studied. The best helix-former is (i+4)E,K, which shows approximately 80% helicity in 0.01 M NaCl at pH 7 and 0 degree C. (ii) The two i+4 peptides show more helix formation than the i+3 peptides. pH titration gives no evidence for helix stabilization by i+3 ion pairs. (iii) Surprisingly, the i+4 peptides form more stable helices than the i+3 peptides at extremes of pH (pH 2 and pH 12) as well as at pH 7. These results may be explained by helix stabilization through Glu-...Lys+ salt bridges at pH 7 and singly charged H bonds at pH 2 (Glu0...Lys+) and pH 12 (Glu-...Lys0). The reason why these links stabilize the alpha-helix more effectively in the i+4 than in the i+3 peptides is not known. (iv) Reversal of the positions of glutamic and lysine residues usually affects helix stability in the manner expected for interaction of these charged groups with the helix dipole. (v) alpha-Helix formation in these alanine-based peptides is enthalpy-driven, as is helix formation by the C-peptide of ribonuclease A.
849 citations
TL;DR: Short, 16-residue, alanine-based peptides show stable alpha-helix formation in H2O, and the likely explanation for these results is that individualAlanine residues have a high helical potential.
Abstract: Short, 16-residue, alanine-based peptides show stable alpha-helix formation in H2O. This result is surprising when contrasted with the classical view that regards the alpha-helix as a marginally stable structure in H2O and considers short helices unstable. The alanine-based peptides are solubilized by insertion of three or more residues of a single charge type, lysine (+) or glutamic acid (-). The results cannot be explained by helix stabilization resulting from concentration-dependent association or by the interaction of charged residues with the helix dipole. Our results are not predicted by the parameters for alanine and lysine that have been determined by the "host-guest" method: these parameters predict that a 16-residue peptide should not show measurable alpha-helix formation. Analysis of the role of the hydrophobic interaction in alpha-helix formation [Richards, F.M. & Richmond, T. (1978) in Molecular Interactions and Activity in Proteins, Ciba Foundation Symposium 60, ed. Wolstenholme, G.E. (Excepta Medica Amsterdam), pp. 23-25] does not show an unusually strong hydrophobic interaction in a helical block of alanine residues. The likely explanation for our results is, therefore, that individual alanine residues have a high helical potential. It is not yet known whether any other amino acids show this property, and the origin of this property is also unknown.
716 citations
638 citations