Transmembrane peptide-induced lipid sorting and mechanism of Lalpha-to-inverted phase transition using coarse-grain molecular dynamics

Molecular dynamics results are presented for a coarse-grain model of 1,2-di-n-alkanoyl-sn-glycero-3-phosphocholine, water, and a capped cylindrical model of a transmembrane peptide. We first demonstrate that different alkanoyl-length lipids are miscible in the liquid-disordered lamellar (Lalpha) phase. The transmembrane peptide is constructed of hydrophobic sites with hydrophilic caps. The hydrophobic length of the peptide is smaller than the hydrophobic thickness of a bilayer consisting of an equal mixture of long and short alkanoyl tail lipids. When incorporated into the membrane, a meniscus forms in the vicinity of the peptide and the surrounding area is enriched in the short lipid. The meniscus region draws water into it. In the regions that are depleted of water, the bilayers can fuse. The lipid headgroups then rearrange to solvate the newly formed water pores, resulting in an inverted phase. This mechanism appears to be a viable pathway for the experimentally observed Lalpha-to-inverse hexagonal (HII) peptide-induced phase transition.     

Hydrophobic mismatch in gramicidin A'/lecithin systems.

Gramicidin A' (GA') has been added to three lipid systems of varying hydrophobic thicknesses: dimyristoyllecithin (DML), dipalmitoyllecithin (DPL), and distearoyllecithin (DSL). The similarity in length between the hydrophobic portion of GA' and the hydrocarbon chains of the lipid bilayers has been studied by using 31P and 2H NMR. Hydrophobic mismatch has been found to be most severe in the DML bilayer system and minimal in the case of DSL. In addition, the effects of hydrophobic mismatch on the cooperative properties of the bilayer have been obtained from 2H NMR relaxation measurements. The results indicate that incorporation of the peptide into the bilayer disrupts the cooperative director fluctuations characteristic of pure multilamellar lipid dispersions. Finally, the GA'/lecithin ratio at which the well-known transformation from bilayer to reverse hexagonal (HII) phase occurs (Van Echteld et al., 1982; Chupin et al., 1987) is shown to depend on the acyl chain length of the phospholipid. A rationale is proposed for this chain length dependence.     

Modulation of polymorphic properties of dielaidoylphosphatidylethanolamine by the antineoplastic ether lipid 1-O-octadecyl-2-O-methyl-glycero-3-phosphocholine

The capacity of the antineoplastic ether lipid 1-O-octadecyl-2-O-methyl-glycero-3-phosphocholine (ET-18-OCH3) to modulate the polymorphic properties of dielaidoylphosphatidylethanolamine has been studied using biophysical techniques. Differential scanning calorimetry showed that ET-18-OCH3 depresses the onset of the Lbeta to Lalpha phase transition, decreasing also DeltaH of the transition. At the same time, the onset of the transition from Lalpha to inverted hexagonal HII phase was gradually increased as the ether lipid concentration was increased, totally disappearing at concentrations higher than 5 mol%. Small-angle X-ray diffraction and 31P-NMR confirmed that ET-18-OCH3 induced that the appearance of the inverted hexagonal HII phase was shifted towards higher temperatures completely disappearing at concentrations higher than 5 mol%. These results were used to elaborate a partial phase diagram and they were discussed as a function of the molecular action of ET-18-OCH3.     

Peptide Nanopores and Lipid Bilayers: Interactions by Coarse-Grained Molecular-Dynamics Simulations

A set of 49 protein nanopore-lipid bilayer systems was explored by means of coarse-grained molecular-dynamics simulations to study the interactions between nanopores and the lipid bilayers in which they are embedded. The seven nanopore species investigated represent the two main structural classes of membrane proteins (α-helical and β-barrel), and the seven different bilayer systems range in thickness from ∼28 to ∼43 Å. The study focuses on the local effects of hydrophobic mismatch between the nanopore and the lipid bilayer. The effects of nanopore insertion on lipid bilayer thickness, the dependence between hydrophobic thickness and the observed nanopore tilt angle, and the local distribution of lipid types around a nanopore in mixed-lipid bilayers are all analyzed. Different behavior for nanopores of similar hydrophobic length but different geometry is observed. The local lipid bilayer perturbation caused by the inserted nanopores suggests possible mechanisms for both lipid bilayer-induced protein sorting and protein-induced lipid sorting. A correlation between smaller lipid bilayer thickness (larger hydrophobic mismatch) and larger nanopore tilt angle is observed and, in the case of larger hydrophobic mismatches, the simulated tilt angle distribution seems to broaden. Furthermore, both nanopore size and key residue types (e.g., tryptophan) seem to influence the level of protein tilt, emphasizing the reciprocal nature of nanopore-lipid bilayer interactions.     

Energetics of hydrophobic matching in lipid-protein interactions

Lipid chain length modulates the activity of transmembrane proteins by mismatch between the hydrophobic span of the protein and that of the lipid membrane. Relative binding affinities of lipids with different chain lengths are used to estimate the excess free energy of lipid-protein interaction that arises from hydrophobic mismatch. For a wide range of integral proteins and peptides, the energy cost is much less than the elastic penalty of fully stretching or compressing the lipid chains to achieve complete hydrophobic matching. The chain length dependences of the free energies of lipid association are described by a model that combines elastic chain extension with a free energy term that depends linearly on the extent of residual mismatch. The excess free energy densities involved lie in the region of 0.5-2.0 k_BT.nm⁻². Values of this size could arise from exposure of hydrophobic groups to polar portions of the lipid or protein, but not directly to water, or alternatively from changes in tilt of the transmembrane helices that are energetically comparable to those activating mechanosensitive channels. The influence of hydrophobic mismatch on dimerization of transmembrane helices and their transfer between lipid vesicles, and on shifts in chain-melting transitions of lipid bilayers by incorporated proteins, is analyzed by using the same thermodynamic model. Segmental order parameters confirm that elastic lipid chain distortions are insufficient to compensate fully for the mismatch, but the dependence on chain length with tryptophan-anchored peptides requires that the free energy density of hydrophobic mismatch should increase with increasing extent of mismatch.     

Influence of vitamin E on phosphatidylethanolamine lipid polymorphism

The effect of vitamin E, in its major form alpha-tocopherol and its synthetic analog alpha-tocopheryl acetate, on phosphatidylethanolamine lipid polymorphism has been studied by mean of differential scanning calorimetry and 31P-nuclear magnetic resonance techniques. From the interaction of these tocopherols with dielaidoylphosphatidylethanolamine it is concluded that both molecules promote the formation of the hexagonal HII phase at temperatures lower than those of the pure phospholipid. When the tocopherols were incorporated in the saturated dimiristoylphosphatidylethanolamine, which has been shown not to undergo bilayer to hexagonal HII phase transition, up to 90 degrees C, they induce the phospholipid to partially organize in hexagonal HII phase. From our experiments it is shown that alpha-tocopherol is more effective than its analog in promoting HII phase in these systems. It is also shown that, while alpha-tocopheryl acetate does not significantly perturb the gel to liquid-crystalline phase transition of dimirystoylphosphatidylethanolamine, alpha-tocopherol does so and more than one peak appears in the calorimetric profile, indicating that lateral phase separations are taking place.     

Temperature and compositional dependence of the structure of hydrated dimyristoyl lecithin.

Differential scanning calorimetry and x-ray diffraction techniques have been used to investigate the structure and phase behavior of hydrated dimyristoyl lecithin (DML) in the hydration range 7.5 to 60 weight % water and the temperature range -10 to +60 degrees C. Four different calorimetric transitions have been observed: T1, a low enthalpy transition (deltaH approximately equal to 1 kcal/mol of DML) at 0 degrees C between lamellar phases (L leads to Lbeta); T2, the low enthalpy "pretransition" at water contents greater than 20 weight % corresponding to the transition Lbeta leads to Pbeta; T3, the hydrocarbon chain order-disorder transition (deltaH = 6 to 7 kcal/mol of DML) representing the transition of the more ordered low temperature phases (Lbeta, Pbeta, or crystal C, depending on the water content) to the lamellar Lalpha phase; T4, a transition occurring at 25--27 degrees C at low water contents representing the transition from the lamellar Lbeta phase to a hydrated crystalline phase C. The structures of the Lbeta, Pbeta, C, and Lalpha phases have been examined as a function of temperature and water content. The Lbeta structure has a lamellar bilayer organization with the hydrocarbon chains fully extended and tilted with respect to the normal to the bilayer plane, but packed in a distorted quasihexagonal lattice. The Pbeta structure consists of lipid bilayer lamellae distorted by a periodic "ripple" in the plane of the lamellae; the hydrocarbon chains are tilted but appear to be packed in a regular hexagonal lattice. The diffraction pattern from the crystalline phase C indexes according to an orthorhombic cell with a = 53.8 A, b = 9.33 A, c = 8.82 A. In the lamellae bilayer Lalpha strucure, the hydrocarbon chains adopt a liquid-like conformation. Analysis of the hydration characteristics and bilayer parameters (lipid thickness, surface area/molecule) of synthetic lecithins permits an evaluation of the generalized hydration and structural behavior of this class of lipids.     

Lipid polymorphism of mixtures of dioleoylphosphatidylethanolamine and saturated and monounsaturated phosphatidylcholines of various chain lengths

The L alpha-HII phase transition behavior of many lipid-water liquid crystals is dominated by the competition between the tendency to curl the lipid layers to an intrinsic radius of curvature and opposing hydrocarbon packing constraints. In particular, packing constraints can increase the free energy of the inverted hexagonal (HII) phase as compared to that of the lamellar (L alpha) phase. This is especially true where the lipid molecule is not long enough to reach into the corners of the lattice in large hexagonal structures necessitated by a large intrinsic radius of curvature. In this paper it is shown that the addition of a minor fraction long-chain lipid to a system of otherwise uniform chain composition can also relax packing constraints, thereby lowering the lamellar to hexagonal transition temperature. For the specific systems used, dioleoylphosphatidylethanolamine (di-18:1c-PE) with minor fractions of 1,2-diacyl-sn-glycero-3-phosphocholines [di-n:1c-PC (n = 14, 18, 22, and 24)], the observed HII lattices systematically increased in size with increasing chain length, suggesting that the chain length also may affect the intrinsic curvature of the mixture. These experiments demonstrate that the lipid "shape concept", which is a qualitative expression of the concept quantitatively described by the intrinsic radius of curvature, is insufficient to understand the L alpha-HII transition. It is necessary to, at least, consider the competition between curvature and packing.     

Tracking phospholipid populations in polymorphism by sideband analyses of 31P magic angle spinning NMR.

A method was developed to track the distributional preferences of phospholipids in polymorphism based on sideband analyses of the 31P magic angle spinning nuclear magnetic resonance spectra. The method was applied to lipid mixtures containing phosphatidylcholine (PtdCho), phosphatidylethanolamine (PtdEtn) and either cholesterol (Chol) or tetradecane, as well as mixtures containing the anionic phosphatidylmethanol, phosphatidylethanolamine, and diolein. The phospholipid composition of coexisting lamellar (Lalpha) and inverted hexagonal (HII) phases remained constant throughout the Lalpha --> HII transition in all mixtures, except those that contained saturated PtdCho and unsaturated PtdEtn in the presence of cholesterol-mixtures that are known to be microimmiscible because of favored associations between Chol and saturated acyl chains. In the latter mixture, saturated PtdCho was enriched in the planar bilayer structure, and unsaturated PtdEtn was enriched in the highly curved HII structure. This enrichment was coincident with an increase in the transition width. When compositional heterogeneity among coexisting phases was observed, it appeared that preexisting lateral microheterogeneities led to compositionally distinct transitional clusters, such that the distributional preferences that resulted were not those of the individual phospholipids.     

Analytical derivation of thermodynamic properties of bilayer membrane with interdigitation

We consider a model of bilayer lipid membrane with interdigitation, in which the lipid tails of the opposite monolayers interpenetrate. The interdigitation is modeled by linking tails of the hydrophobic chains in the opposite monolayers within bilayer as a first approximation. This model corresponds to the types of interdigitation that are not related with the areal “hydrophobic” dilation of the membrane. A number of essential thermodynamical characteristics are calculated analytically and compared with the ones of a regular bilayer membrane without interdigitation. Important difference between lateral pressure profiles at the layers interface for linked and regular bilayer models is found. In the linked case, the lateral pressure mid-plane peak disappears, while the entropy decreases and the free energy per chain increases. Within our model we found that in case of elongation of the chains inside a nucleus of, e.g., liquid-condensed phase, homogeneous interdigitation would be more costly for the membrane’s free energy than energy of the hydrophobic mismatch between the elongated chains and the liquid-expanded surrounding. Nonetheless, an inhomogeneous interdigitation along the nucleus boundary may occur inside a “belt” of a width that varies approximately with the hydrophobic mismatch amplitude.     

Spin-label characterisation of the lamellar-to-hexagonal (HII) phase transition in egg phosphatidylethanolamine aqueous dispersions

The thermotropic behaviour of egg yolk phosphatidylethanolamine dispersions in excess aqueous phase has been investigated by spin label electron spin resonance spectroscopy and differential thermal analysis. Phosphatidylethanolamine isomers spin-labelled at six different positions along the acyl chain, and steroid spin labels, indicate both gel-fluid lamellar and lamellar-reverse hexagonal (HII) phase transitions, in agreement with complementary calorimetric studies. Analysis of spin label data shows that the transition to the HII phase is accompanied by an increase in conformational freedom of the acyl chain, more pronounced towards the methyl terminus, and representing an increase in the population of gauche isomers which can only be accommodated by a transition to the non-bilayer phase. Raising the bulk pH to, and above, pH 8.5 results in stabilisation of the bilayer phase and no transition to the HII phase is observed. The phosphatidylethanolamine spin labels also indicate a polarity profile which is characteristic of each phase.     

Modulation of the bilayer to hexagonal phase transition of phosphatidylethanolamines by acylglycerols

The effect of mono-, di- and triacylglycerols on the bilayer to hexagonal phase (HII) transition was studied by differential scanning calorimetry and 31P-NMR spectroscopy. The acylglycerols were mixed with either dielaidoylphosphatidylethanoline or with 1-palmitoyl-2-oleoylphosphatidylethanolamine. Acylglycerols of lauric, oleic and stearic acids were utilized. All of the acylglycerols lowered the bilayer to HII phase transition temperature. Diacylglycerols were much better HII phase promoters than monoacylglycerols while triacylglycerols were the most potent bilayer phase destabilizers. Fatty acid composition generally had less of an effect except for the monoacylglycerols where bilayer destabilization increased from monolaurin to monostearin to monoolein. The most marked difference in behaviour resulting from changes in the fatty acid composition of the acylglycerol occurred with tristearin. This was the only acylglycerol which decreased the bilayer to HII phase transition temperature only below a mol fraction of 0.005. Above this mol fraction, further addition of tristearin had no effect on the bilayer to HII phase transition. These results suggest that the tristearin has limited solubility in phosphatidylethanolamine.     

N-succinyldioleoylphosphatidylethanolamine: structural preferences in pure and mixed model membranes

The structural preferences of the pH-sensitive phospholipid, N-succinyldioleoylphosphatidylethanolamine (N-succinyl-DOPE), have been examined alone and in mixtures with DOPE by 31P-NMR, fluorescence energy transfer, and freeze-fracture techniques. The basic polymorphic behavior of pure N-succinyl-DOPE and DOPE/N-succinyl-DOPE lipid systems and the influence of calcium and pH were investigated. It is shown that, similar to other negatively charged acidic phospholipids, N-succinyl-DOPE adopts the bilayer organization upon hydration. This structure is maintained at both pH 7.4 and 4.0 in the presence or absence of calcium. In the mixed lipid system, N-succinyl-DOPE can stabilize the non-bilayer lipid, DOPE, into a bilayer structure at both pH 7.4 and 4.0 at more than 10 mol% N-succinyl-DOPE, although a narrow 31P-NMR lineshape is observed at acidic pH values. This corresponds to the presence of smaller vesicles as shown by quasi-elastic light scattering measurements. Addition of equimolar calcium (with respect to N-succinyl-DOPE) to the DOPE/N-succinyl-DOPE systems induces the hexagonal HII phase at both pH values. In unilamellar systems with similar lipid composition the addition of Ca2+ results in membrane fusion as indicated by fluorescence energy-transfer experiments. These findings are discussed with regard to the molecular mechanism of the bilayer to hexagonal HII phase transition and membrane fusion and the utility of N-succinyl-DOPE containing pH-sensitive vesicles as drug-delivery vehicles.     

Simulation studies of protein-induced bilayer deformations, and lipid-induced protein tilting, on a mesoscopic model for lipid bilayers with embedded proteins

Biological membranes are complex and highly cooperative structures. To relate biomembrane structure to their biological function it is often necessary to consider simpler systems. Lipid bilayers composed of one or two lipid species, and with embedded proteins, provide a model system for biological membranes. Here we present a mesoscopic model for lipid bilayers with embedded proteins, which we have studied with the help of the dissipative particle dynamics simulation technique. Because hydrophobic matching is believed to be one of the main physical mechanisms regulating lipid-protein interactions in membranes, we considered proteins of different hydrophobic length (as well as different sizes). We studied the cooperative behavior of the lipid-protein system at mesoscopic time- and lengthscales. In particular, we correlated in a systematic way the protein-induced bilayer perturbation, and the lipid-induced protein tilt, with the hydrophobic mismatch (positive and negative) between the protein hydrophobic length and the pure lipid bilayer hydrophobic thickness. The protein-induced bilayer perturbation was quantified in terms of a coherence length, xi(P), of the lipid bilayer hydrophobic thickness profile around the protein. The dependence on temperature of xi(P), and the protein tilt-angle, were studied above the main-transition temperature of the pure system, i.e., in the fluid phase. We found that xi(P) depends on mismatch, i.e., the higher the mismatch is, the longer xi(P) becomes, at least for positive values of mismatch; a dependence on the protein size appears as well. In the case of large model proteins experiencing extreme mismatch conditions, in the region next to the so-called lipid annulus, there appears an undershooting (or overshooting) region where the bilayer hydrophobic thickness is locally lower (or higher) than in the unperturbed bilayer, depending on whether the protein hydrophobic length is longer (or shorter) than the pure lipid bilayer hydrophobic thickness. Proteins may tilt when embedded in a too-thin bilayer. Our simulation data suggest that, when the embedded protein has a small size, the main mechanism to compensate for a large hydrophobic mismatch is the tilt, whereas large proteins react to negative mismatch by causing an increase of the hydrophobic thickness of the nearby bilayer. Furthermore, for the case of small, peptidelike proteins, we found the same type of functional dependence of the protein tilt-angle on mismatch, as was recently detected by fluorescence spectroscopy measurements.     

Diacylglycerols, lysolecithin, or hydrocarbons markedly alter the bilayer to hexagonal phase transition temperature of phosphatidylethanolamines

The bilayer to hexagonal phase transition temperatures of dielaidoylphosphatidylethanolamine and 1-palmitoyl-2-oleoylphosphatidylethanolamine are 65.6 and 71.4 degrees C, respectively. Using high-sensitivity differential scanning calorimetry, I have shown that these transition temperatures are extremely sensitive to the presence of small amounts of other lipid components. For example, at a mole fraction of only 0.01, dilinolenin lowers the bilayer to hexagonal phase transition temperature of 1-palmitoyl-2-oleoyl-phosphatidylethanolamine by 8.5 degrees C. Other diacylglycerols have similar effects on this transition temperature, although the degree of unsaturation of the acyl chains has some effect, with distearin being less potent. In comparison, the 20-carbon alkane eicosane lowers this transition temperature by 5 degrees C, while palmitoyl-lysolecithin raises it by 2.5 degrees C. Similar effects of these additives on the bilayer to to hexagonal phase transition temperature are observed with dielaidoylphosphatidylethanolamine. At these concentrations of additive, there is no effect on the gel-state to liquid-crystalline-state transition temperature. The observed shifts in the temperature of the bilayer to the hexagonal phase transition can be qualitatively interpreted in terms of the effects of these additives on the hydrophilic surface area and on the hydrophobic volume. Substances expanding the hydrophobic domain promote hexagonal phase formation and lower the bilayer to hexagonal phase transition temperature. The sensitivity of the bilayer to hexagonal phase transition temperature to the presence of additives is at least as great as that which has been observed for any other lipid phase transition.     

Spin-labeled gramicidin a: channel formation and dissociation

Gramicidin A was studied by continuous wave electron spin resonance (CW-ESR) and by double-quantum coherence electron spin resonance (DQC-ESR) in several lipid membranes (using samples that were macroscopically aligned by isopotential spin-dry ultracentrifugation) and vesicles. As a reporter group, the nitroxide spin-label was attached at the C-terminus yielding the spin-labeled product (GAsl). ESR spectra of aligned membranes containing GAsl show strong orientation dependence. In DPPC and DSPC membranes at room temperature the spectral shape is consistent with high ordering, which, in conjunction with the observed high polarity of the environment of the nitroxide, is interpreted in terms of the nitroxide moiety being close to the membrane surface. In contrast, spectra of GAsl in DMPC membranes indicate deeper embedding and tilt of the NO group. The GAsl spectrum in the DPPC membrane at 35 degrees C (the gel to Pbeta phase transition) exhibits sharp changes, and above this temperature becomes similar to that of DMPC. The dipolar spectrum from DQC-ESR clearly indicates the presence of pairs in DMPC membranes. This is not the case for DPPC, rapidly frozen from the gel phase; however, there are hints of aggregation. The interspin distance in the pairs is 30.9 A, in good agreement with estimates for the head-to-head GAsl dimer (the channel-forming conformation), which matches the hydrophobic thickness of the DMPC bilayer. Both DPPC and DSPC, apparently as a result of hydrophobic mismatch between the dimer length and bilayer thickness, do not favor the channel formation in the gel phase. In the Pbeta and Lalpha phases of DPPC (above 35 degrees C) the channel dimer forms, as evidenced by the DQC-ESR dipolar spectrum after rapid freezing. It is associated with a lateral expansion of lipid molecules and a concomitant decrease in bilayer thickness, which reduces the hydrophobic mismatch. A comparison with studies of dimer formation by other physical techniques indicates the desirability of using low concentrations of GA (approximately 0.4-1 mol %) accessible to the ESR methods employed in the study, since this yields non-interacting dimer channels.     

Transmembrane helices of membrane proteins may flex to satisfy hydrophobic mismatch

A novel mechanism for membrane modulation of transmembrane protein structure, and consequently function, is suggested in which mismatch between the hydrophobic surface of the protein and the hydrophobic interior of the lipid bilayer induces a flexing or bending of a transmembrane segment of the protein. Studies on model hydrophobic transmembrane peptides predict that helices tilt to submerge the hydrophobic surface within the lipid bilayer to satisfy the hydrophobic effect if the helix length exceeds the bilayer width. The hydrophobic surface of transmembrane helix 1 (TM1) of lactose permease, LacY, is accessible to the bilayer, and too long to be accommodated in the hydrophobic portion of a typical lipid bilayer if oriented perpendicular to the membrane surface. Hence, nuclear magnetic resonance (NMR) data and molecular dynamics simulations show that TM1 from LacY may flex as well as tilt to satisfy the hydrophobic mismatch with the bilayer. In an analogous study of the hydrophobic mismatch of TM7 of bovine rhodopsin, similar flexing of the transmembrane segment near the conserved NPxxY sequence is observed. As a control, NMR data on TM5 of lacY, which is much shorter than TM1, show that TM5 is likely to tilt, but not flex, consistent with the close match between the extent of hydrophobic surface of the peptide and the hydrophobic thickness of the bilayer. These data suggest mechanisms by which the lipid bilayer in which the protein is embedded modulates conformation, and thus function, of integral membrane proteins through interactions with the hydrophobic transmembrane helices.     

Transmembrane helices of membrane proteins may flex to satisfy hydrophobic mismatch

A novel mechanism for membrane modulation of transmembrane protein structure, and consequently function, is suggested in which mismatch between the hydrophobic surface of the protein and the hydrophobic interior of the lipid bilayer induces a flexing or bending of a transmembrane segment of the protein. Studies on model hydrophobic transmembrane peptides predict that helices tilt to submerge the hydrophobic surface within the lipid bilayer to satisfy the hydrophobic effect if the helix length exceeds the bilayer width. The hydrophobic surface of transmembrane helix 1 (TM1) of lactose permease, LacY, is accessible to the bilayer, and too long to be accommodated in the hydrophobic portion of a typical lipid bilayer if oriented perpendicular to the membrane surface. Hence, nuclear magnetic resonance (NMR) data and molecular dynamics simulations show that TM1 from LacY may flex as well as tilt to satisfy the hydrophobic mismatch with the bilayer. In an analogous study of the hydrophobic mismatch of TM7 of bovine rhodopsin, similar flexing of the transmembrane segment near the conserved NPxxY sequence is observed. As a control, NMR data on TM5 of lacY, which is much shorter than TM1, show that TM5 is likely to tilt, but not flex, consistent with the close match between the extent of hydrophobic surface of the peptide and the hydrophobic thickness of the bilayer. These data suggest mechanisms by which the lipid bilayer in which the protein is embedded modulates conformation, and thus function, of integral membrane proteins through interactions with the hydrophobic transmembrane helices.     

Interaction of a peptide model of a hydrophobic transmembrane alpha-helical segment of a membrane protein with phosphatidylcholine bilayers: differential scanning calorimetric and FTIR spectroscopic studies.

High-sensitivity differential scanning calorimetry (DSC) and Fourier transform infrared (FTIR) spectroscopy were used to study the interaction of a synthetic model hydrophobic peptide, Lys2-Gly-Leu24-Lys2-Ala-amide, and members of the homologous series of n-saturated diacylphosphatidylcholines. In the low range of peptide mole fractions, the DSC thermograms exhibited by the lipid/peptide mixtures are resolvable into two components. One of these components is fairly narrow, highly cooperative, and exhibits properties which are similar to but not identical with those of the pure lipid. In addition, the fractional contribution of this component to the total enthalpy change, the peak transition temperature, and cooperativity decrease with an increase in peptide concentration, more or less independently of acyl chain length. The other component is very broad and predominates in the high range of peptide concentration. These two components have been assigned to the chain-melting phase transitions of populations of bulk lipid and peptide-associated lipid, respectively. Moreover, when the mean hydrophobic thickness of the PC bilayer is less than the peptide hydrophobic length, the peptide-associated lipid melts at higher temperatures than does the bulk lipid and vice versa. In addition, the chain-melting enthalpy of the broad endotherm does not decrease to zero even at high peptide concentrations, suggesting that this peptide reduces but do not abolish the cooperative gel/liquid-crystalline phase transition of the lipids with which it is in contact. Our DSC results indicate that the width of the phase transition observed at high peptide concentration is inversely but discontinuously related to hydrocarbon chain length and that gel phase immiscibility occurs when the hydrophobic thickness of the bilayer greatly exceeds the hydrophobic length of the peptide. The FTIR spectroscopic data indicate that the peptide forms a very stable alpha-helix under all of our experimental conditions but that small distortions of its alpha-helical conformation are induced in response to any mismatch between peptide hydrophobic length and bilayer hydrophobic thickness. These results also indicate that the peptide alters the conformational disposition of the acyl chains in contact with it and that the resultant conformational changes in the lipid hydrocarbon chains tend to minimize the extent of mismatch of peptide hydrophobic length and bilayer hydrophobic thickness.     

The interaction of coenzyme Q with phosphatidylethanolamine membranes.

Coenzyme Q (CoQ) is a component of the mitochondrial respiratory chain which carries out additional membrane functions, such as acting as an antioxidant. The location of CoQ in the membrane and the interaction with the phospholipid bilayer is still a subject of debate. The interaction of CoQ in the oxidized (ubiquinone-10) and reduced (ubiquinol-10) state with membrane model systems of 1,2-dielaidoyl-sn-glycero-3-phosphoethanolamine (Ela2Gro-P-Etn) has been studied by means of differential scanning calorimetry (DSC), 31P-nuclear magnetic resonance (31P-NMR) and small angle X-ray diffraction (SAXD). Ubiquinone-10 did not visibly affect the lamellar gel to lamellar liquid-crystalline phase transition of Ela2Gro-P-Etn, but it clearly perturbed the multicomponent lamellar liquid-crystalline to lamellar gel phase transition of the phospholipid. The perturbation of both transitions was more effective in the presence of ubiquinol-10. A location of CoQ forming head to head aggregates in the center of the Ela2Gro-P-Etn bilayer with the polar rings protruding toward the phospholipid acyl chains is suggested. The formation of such aggregates are compatible with the strong hexagonal HII phase promotion ability found for CoQ. This ability was evidenced by the shifting of the lamellar to hexagonal HII phase transition to lower temperatures and by the appearance of the characteristic hexagonal HII 31P-NMR resonance and SAXD pattern at temperatures at which the pure Ela2Gro-P-Etn is still organized in extended bilayer structures. The influence of CoQ on the thermotropic properties and phase behavior of Ela2Gro-P-Etn is discussed in relation to the role of CoQ in the membrane.