Investigating the conformational coupling between the transmembrane and cytoplasmic domains of a single-spanning membrane protein. A 1H-NMR study.

PMP1 is a 38-residue single-spanning membrane protein whose C-terminal cytoplasmic domain, Y25-F38, is highly positively charged. The conformational coupling between the transmembrane span and the cytoplasmic domain of PMP1 was investigated from 1H-nuclear magnetic resonance data of two synthetic fragments: F9-F38, i.e. 80% of the whole sequence, and Y25-F38, the isolated cytoplasmic domain. Highly disordered in aqueous solution, the Y25-F38 peptide adopts a well-defined conformation in the presence of dodecylphosphocholine micelles. Compared with the long PMP1 fragment, this structure exhibits both native and non-native elements. Our results make it possible to assess the influence of a hydrophobic anchor on the intrinsic conformational propensity of a cytoplasmic domain.

Concerted influence of key amino acids on the lipid binding properties of a single-spanning membrane protein: NMR and mutational analysis.

Finding the combinations of key amino acids involved in the interaction network underlying the interfacial features of membrane proteins would contribute to a better understanding of their sequence-structure-function relationships and the role of anionic phospholipids. To further address these questions, we performed mutational analysis associated with NMR experiments on synthetic fragments of the single-spanning membrane protein PMP1 that exhibit binding specificity for phosphatidylserine (PS). The aromatic and glutamine residues of the helix part of the PMP1 cytoplasmic domain were mutated. (1)H NMR experiments were carried out using perdeuterated DPC micelles as a membrane-like environment, in the absence and presence of small amounts of either POPC or POPS lipids. From intermolecular NOEs and chemical shift data, specific and nonspecific aspects of peptide-phospholipid interactions were distinguished. The major finding of our study is to reveal the concerted influence of a tryptophan and a glutamine residue on the interfacial conformation and lipid binding specificity of the PMP1 cytoplasmic domain.

15N NMR relaxation as a probe for helical intrinsic propensity: the case of the unfolded D2 domain of annexin I.

The isolated D2 domain of annexin I is unable to adopt a tertiary fold but exhibits both native and non-native residual structures. It thus constitutes an attractive model for the investigation of dynamics of partially folded states in the context of protein folding and stability. 15N relaxation parameters of the D2 domain have been acquired at three different magnetic fields, 500, 600 and 800 MHz. This enables the estimation of the contribution of conformational exchange to the relaxation parameters on the micro- to millisecond time scale, thus providing a suitable data set for the description of motions on the pico- and nanosecond time scale. The analysis of the seven spectral densities obtained (J(0), J(50 MHz), J(60 MHz), J(80 MHz), , , ) provides complementary and meaningful results on the conformational features of the D2 domain structure previously depicted by chemical shift and NOE data. Especially, residual helix segments exhibit distinct dynamical behaviors that are related to their intrinsic helical propensity. Beside the spectral density analysis, a series of models derived from the Lipari and Szabo model-free approach are investigated. Two models containing three parameters are able to reproduce equally well the experimental data within experimental errors but provide different values of order parameters and correlation times. The inability to find a unique model to describe the data emphasizes the difficulty to use and interpret the model-free parameters in the case of partially or fully unfolded proteins consisting of a wide range of interconverting conformers.

PMP1 18-38, a yeast plasma membrane protein fragment, binds phosphatidylserine from bilayer mixtures with phosphatidylcholine: a (2)H-NMR study.

PMP1 is a 38-residue plasma membrane protein of the yeast Saccharomyces cerevisiae that regulates the activity of the H(+)-ATPase. The cytoplasmic domain conformation results in a specific interfacial distribution of five basic side chains, thought to strongly interact with anionic phospholipids. We have used the PMP1 18-38 fragment to carry out a deuterium nuclear magnetic resonance ((2)H-NMR) study for investigating the interactions between the PMP1 cytoplasmic domain and phosphatidylserines. For this purpose, mixed bilayers of 1-palmitoyl, 2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl, 2-oleoyl-sn-glycero-3-phosphoserine (POPS) were used as model membranes (POPC/POPS 5:1, m/m). Spectra of headgroup- and chain-deuterated POPC and POPS phospholipids, POPC-d4, POPC-d31, POPS-d3, and POPS-d31, were recorded at different temperatures and for various concentrations of the PMP1 fragment. Data obtained from POPS deuterons revealed the formation of specific peptide-POPS complexes giving rise to a slow exchange between free and bound PS lipids, scarcely observed in solid-state NMR studies of lipid-peptide/protein interactions. The stoichiometry of the complex (8 POPS per peptide) was determined and its significance is discussed. The data obtained with headgroup-deuterated POPC were rationalized with a model that integrates the electrostatic perturbation induced by the cationic peptide on the negatively charged membrane interface, and a "spacer" effect due to the intercalation of POPS/PMP1f complexes between choline headgroups.

NMR conformational study of the sixth transmembrane segment of sarcoplasmic reticulum Ca2+-ATPase.

In current topological models, the sarcoplasmic reticulum Ca2+-ATPase contains 10 putative transmembrane spans (M1-M10), with spans M4/M5/M6 and probably M8 participating in the formation of the membranous calcium-binding sites. We describe here the conformational properties of a synthetic peptide fragment (E785-N810) encompassing the sixth transmembrane span (M6) of Ca2+-ATPase. Peptide M6 includes three residues (N796, T799, and D800) out of the six membranous residues critically involved in the ATPase calcium-binding sites. 2D-NMR experiments were performed on the M6 peptide selectively labeled with 15N and solubilized in dodecylphosphocholine micelles to mimic a membrane-like environment. Under these conditions, M6 adopts a helical structure in its N-terminal part, between residues I788 and T799, while its C-terminal part (G801-N810) remains disordered. Addition of 20% trifluoroethanol stabilizes the alpha-helical N-terminal segment of the peptide, and reveals the propensity of the C-terminal segment (G801-L807) to form also a helix. This second helix is located at the interface or in the aqueous environment outside the micelles, while the N-terminal helix is buried in the hydrophobic core of the micelles. Furthermore, the two helical segments of M6 are linked by a flexible hinge region containing residues T799 and D800. These conformational features may be related to the transient formation of a Schellman motif (L797VTDGL802) encoded in the M6 sequence, which probably acts as a C-cap of the N-terminal helix and induces a bend with respect to the helix axis. We propose a model illustrating two conformations of M6 and its insertion in the membrane. The presence of a flexible region within M6 would greatly facilitate concomitant participation of all three residues (N796, T799, and D800) believed to be involved in calcium complexation.

Dodecylphosphocholine micelles as a membrane-like environment: new results from NMR relaxation and paramagnetic relaxation enhancement analysis.

To further examine to what extent a dodecyl-phosphocholine (DPC) micelle mimics a phosphatidylcholine bilayer environment, we performed 13C, 2H, and 31P NMR relaxation measurements. Our data show that the dynamic behavior of DPC phosphocholine groups at low temperature (12 degrees C) corresponds to that of a phosphatidylcholine interface at high temperature (51 degrees C). In the presence of helical peptides, a PMP1 fragment, or an annexin fragment, the DPC local dynamics are not affected whereas the DPC aggregation number is increased to match an appropriate area/volume ratio for accommodating the bound peptides. We also show that quantitative measurements of paramagnetic relaxation enhancements induced by small amounts of spin-labeled phospholipids on peptide proton signals provide a meaningful insight on the location of both PMP1 and annexin fragments in DPC micelles. The paramagnetic contributions to the relaxation were extracted from intra-residue cross-peaks of NOESY spectra for both peptides. The location of each peptide in the micelles was found consistent with the corresponding relaxation data. As illustrated by the study of the PMP1 fragment, paramagnetic relaxation data also allow us to supply the missing medium-range NOEs and therefore to complete a standard conformational analysis of peptides in micelles.

1H- and 2H-NMR studies of a fragment of PMP1, a regulatory subunit associated with the yeast plasma membrane H(+)-ATPase. Conformational properties and lipid-peptide interactions.

PMP1 is a 38-residue polypeptide associated with the yeast plasma membrane H(+)-ATPase, found to regulate the enzyme activity. To investigate the molecular basis of the PMP1 biological function, the conformational properties of a synthetic PMP1 fragment, A18-F38, comprising the predicted C-terminal cytoplasmic domain and a part of the transmembrane anchor have been studied by 1H- and 2H-NMR spectroscopies. High resolution 1H-NMR experiments showed that, in deuterated DPC micelles, the A18-G34 segment adopts a well defined helix conformation. Our data suggest that the whole PMP1 molecule forms a unique helix whose axis might be slightly tilted with respect to the bilayer normal. Protonated DPC, DMPC and DMPS were incorporated in deuterated micelles containing the PMP1 fragment for studying lipid-peptide interactions. Unusually strong and selective intermolecular NOEs between lipid chain and peptide side chain protons, especially those of the unique Trp residue, were observed. Solid state 2H-NMR experiments performed on pure deuterated POPC and mixed deuterated POPC:POPS (5:1) bilayers revealed that the PMP1 fragment specifically interacts with negatively charged PS lipids.

Expression, purification, and characterization of Sss1p, an essential component of the yeast Sec61p protein translocation complex.

Sss1p, a 8.9-kDa membrane protein, is an essential component of the protein translocation complex involved in the transport of secretory proteins across the Saccharomyces cerevisiae endoplasmic reticulum membrane. In order to determine the high resolution structure of Sss1p by NMR, we have undertaken its overexpression and purification. We first inserted the yeast SSS1 gene into the pGEX-2T plasmid expression vector. Sss1p was expressed as fusions with Schistosoma japonica glutathione S-transferase (GST-Sss1p) in MC1061 Escherichia coli cells. Maximum yield of GST-Sss1p was obtained from cells harvested 2 h after induction at 37 degreesC in Luria broth medium. GST-Sss1p was found associated predominantly with the membrane pool and was readily extracted with Triton X-100. Detergent-solubilized GST-Sss1p was isolated by adsorption on glutathione-agarose beads. Sss1p was released from its GST carrier by cleavage with thrombin and its recovery was maximized by addition of dodecyl maltoside. Desorbed Sss1p was loaded on a high-performance liquid chromatography hydroxyapatite column equilibrated in phosphate buffer supplemented with dodecyl maltoside and the fractions containing Sss1p were subsequently purified to homogeneity by reverse-phase chromatography on a C4 column. The entire purification protocol can be completed in 5-6 h and yields about 0.4 mg of Sss1p per gram of transformed cells. CD and preliminary 1H NMR experiments show that purified Sss1p solubilized in SDS micelles is very stable and adopts a helical secondary structure.

A conformational equilibrium in a protein fragment caused by two consecutive capping boxes: 1H-, 13C-NMR, and mutational analysis.

The conformational properties of an 18 residues peptide spanning the entire sequence, L1KTPA5QFDAD10ELRAA15MKG, of the first helix (A-helix) of domain 2 of annexin I, were thoroughly investigated. This fragment exhibits several singular features, and in particular, two successive potential capping boxes, T3xxQ6 and D8xxE11. The former corresponds to the native hydrogen bond network stabilizing the alpha helix N-terminus in the protein; the latter is a non-native capping box able to break the helix at residue D8, and is observed in the domain 2 partially folded state. Using 2D-NMR techniques, we showed that two main populations of conformers coexist in aqueous solution. The first corresponds to a single helix extending from T3 to K17. The second corresponds to a broken helix at residue Ds. Four mutants, T3A, F7A, D8A, and E11A, were designed to further analyze the role of key amino acids in the equilibrium between the two ensembles of conformers. The sensitivity of NMR parameters to account for the variations in the populations of conformers was evaluated for each peptide. Our data show the delta13Calpha chemical shift to be the most relevant parameter. We used it to estimate the population ratio in the various peptides between the two main ensembles of conformers, the full helix and the broken helix. For the WT, E11A, and F7A peptides, these ratios are respectively 35/65, 60/40, 60/40. Our results were compared to the data obtained from helix/coil transition algorithms.

Exploring the folding pathways of annexin I, a multidomain protein. I. non-native structures stabilize the partially folded state of the isolated domain 2 of annexin I.

Proteins of the annexin family constitute very attractive models because of their four approximately 70 residue domains, D1 to D4, exhibiting an identical topology comprising five helix segments with only a limited sequence homology of approximately 30%. We focus on the isolated D2 domain, which is only partially folded. A detailed analysis of this equilibrium partially folded state in aqueous solution and micellar solution using 15N-1H multidimensional NMR is presented. Comparison of the residual structure of the entire domain with that of shorter fragments indicates the presence of long-range transient hydrophobic interactions that slightly stabilize the secondary structure elements. The unfolded domain tends to behave as a four-helix, rather than as a five-helix domain. The ensemble of residual structures comprises: (i) a set of native structures consisting of three regions with large helix populations, in rather sharp correspondence with A, B and E helices, and a small helix population in the second part of the C helix; (ii) a set of non-native local structures corresponding to turn-like structures stabilized by several side-chain to side-chain interactions and helix-disruptive side-chains to backbone interactions. Remarkably, residues involved in these local non-native interactions are also involved, in the native structure, in structurally important non-local interactions. During the folding process of annexin I, the local non-native interactions have to switch to native long-range interactions. This structural switch reveals the existence of a sequence-encoded regulation of the folding pathways and kinetics, and emphasizes the key role of the non-native local structures in this regulation.

Exploring the folding pathways of annexin I, a multidomain protein. II. Hierarchy in domain folding propensities may govern the folding process.

In the context of exploring the relationship between sequence and folding pathways, the multi-domain proteins of the annexin family constitute very attractive models. They are constituted of four approximately 70-residue domains, named D1 to D4, with identical topologies but only limited sequence homology of approximately 30%. The domains are organized in a pseudochiral circular arrangement. Here, we report on the folding propensity of the D1 domain of annexin I obtained from overexpression in Escherichia coli. Unlike the D2 domain, which is only partially folded, the isolated D1 domain exhibits autonomous refolding in pure aqueous solution. Similarly, the D3 domain and D2-D3 module were obtained from expression in E. coli but were found to be largely unfolded. No conclusion could be drawn for the D4 domain because it was not possible to extract it from the bacterial inclusion bodies. The data allow us to propose a plausible scenario for the annexin I folding. This working model states that firstly the D1 domain folds, and the D2 and D3 domains remain partly unfolded, facilitating the docking of the D4 domain to the D1 domain. In a second step, the D1 and D4 domains dock, and D4 may fold if already not folded. The final step starts with the stabilization of the D1-D4 module. This stabilization is crucial for allowing the non-native local interactions inside the still partially unfolded D2 domain to switch to the native long-range interactions involving D4. This switch allows the complete folding of D2 and D3. The model proposes a sequential and hierarchical process for the folding of annexin I and emphasizes the role of both native framework and non-native structures in the process.

Effect of (13)C-, (18)O- and (2)H-labeling on the infrared modes of UV-induced phenoxyl radicals.

The structure and environment of redox active tyrosines present in several metalloenzymes can be studied by resonance Raman spectroscopy or Fourier transform infrared difference spectroscopy. Assignments of the vibrational modes in vivo often requires in vitro studies on model compounds. This approach is briefly reviewed. New results are shown on the influence of isotope-labeling on the infrared spectra of tyrosine, [Formula: see text] and phenol radicals obtained in vitro by UV-irradiation. The infrared spectra of the radicals are dominated by the [Formula: see text] mode at 1515-1504 cm(-1). The frequency shifts induced on this mode by (13)C- (2)H-, and (18)O-labeling are reported.

Structure of human annexin I: comparison of homology modelling and crystallographic experiment.

A model of domain II of annexin I has been built by homology modelling using an annexin V crystal structure as a template. The method used is based on that of Summers and Karplus (J Mol Biol (1989) 210, 785-811) and involves the calculation of torsion-angle rotational energy maps to position side chains. The RMS deviation of the backbone heavy atoms between the model and a crystal structure of annexin I is 1.1 A. Similarities and differences in the experimental and model-derived side-chain rotameric conformations and hydrogen-bonding interactions are examined. It is found that whereas many of the side chains are well positioned some of those placed using the 'entropy argument' in which the broadest of the available minima are preferred, are erroneous. The domain is subjected to molecular dynamics simulation in explicit solvent. The simulations are found to 'correct' some of the side-chain rotamer positions that were poorly placed in the homology modelling. Considerable helix instability is seen in the simulations, consistent with the requirement of domain interactions for the structural integrity of the protein.

NMR conformational study of the cytoplasmic domain of the canine Sec61 gamma protein from the protein translocation pore of the endoplasmic reticulum membrane.

Conformational studies of the synthesized N-terminal cytoplasmic domain of the canine Sec61 gamma protein, an essential protein from the translocation pore of secretory proteins across the endoplasmic reticulum membrane, were performed using two-dimensional proton NMR spectroscopy. This canine domain is one of the smallest domains within the homologous protein family and may thus constitute the minimal functional structure. The peptide was solubilized in pure aqueous solution or in the presence of dodecylphosphocholine micelles mimicking a membrane-solution interface. In pure aqueous solution, the peptide is remarkably unfolded. Forming a stable complex with dodecylphosphocholine micelles, it acquires a well-defined alpha-helix-loop-alpha-helix secondary structure, with the helix, highly amphipathic, lying at the micelle surface. The loop comprising four residues is delimited by two flanking helix-capping structures, highly conserved in the whole homologous protein family. No tertiary structure, which could have been revealed by interhelix NOE contacts, was observed. From these experimental results and using general arguments based on sequence information and knowledge of peptide-membrane interactions, a structure of the entire Sec61 gamma protein in membrane bilayers is proposed.

Folding properties of an annexin I domain: a 1H-15N NMR and CD study.

The annexin fold consists of four 70-residue domains with markedly homologous sequences and nearly identical structures. Each domain contains five helices designated A to E. Domain 2 of annexin I was obtained by chemical synthesis including ten specifically labeled residues and studied by 1H-15N NMR and circular dichroism (CD). In pure aqueous solution this annexin domain presents, at most, 25% of residual helix secondary structure compared to 75%-85% for the native helix content and thus does not constitute an autonomous folding unit. Dodecylphosphocholine (DPC) micelles were used to provide the annexin domain with non-specific hydrophobic interactions. The structuring effect of micelles was thoroughly investigated by CD and 1H-15N NMR. Most, but not all, of the native helix secondary structure was recovered at DPC saturation. NMR data made it possible to determine the intrinsic helix propensity hierarchy of the different helix segments of the domain: A approximately B approximately E > C, D. This hierarchy is remarkably well correlated with the location of the helices in the native protein since A, B, and E helices are those in contact with the remaining parts of the protein. This result tends to support the view that, for large proteins like annexins (35 kDa), high intrinsic secondary structure propensities, at least helix propensity, in selected protein segments is necessary for a correct folding process. As a consequence this also indicates that important information concerning the folding pathway is encoded in the protein sequence.

1H and 15N NMR sequential assignment, secondary structure, and tertiary fold of [2Fe-2S] ferredoxin from Synechocystis sp. PCC 6803.

The [2Fe-2S] ferredoxin extracted from Synechocystis sp. PCC 6803 was studied by 1H and 15N nuclear magnetic resonance. Sequence-specific 1H and 15N assignment of amino acid residues far from the paramagnetic cluster (distance higher than 8 A) was performed. Interresidue NOE constraints have allowed the identification of several secondary structure elements: one beta sheet composed of four beta strands, one alpha helix, and two alpha helix turns. The analysis of interresidue NOEs suggests the existence of a disulfide bridge between the cysteine residues 18 and 85. Such a disulfide bridge has never been observed in plant-type ferredoxins. Structure modeling using the X-PLOR program was performed with or without assuming the existence of a disulfide bridge. As a result, two structure families were obtained with rms deviations of 2.2 A. Due to the lack of NOE connectivities resulting from the paramagnetic effect from the [2Fe-2S] cluster, the structures were not well resolved in the region surrounding the [2Fe-2S] cluster, at both extremities of the alpha helix and the C and N terminus segments. In contrast, when taken separately, the beta sheet and the alpha helix were well defined. This work is the first report of a structure model of a plant-type [2Fe-2S] Fd in solution.

Nonnative capping structure initiates helix folding in an annexin I fragment. A 1H NMR conformational study.

A 21-residue peptide, P1AQFD5ADELR10AAMKG15LGTDE20D, corresponding to the (helix A)-loop motif of the second repeat of human annexin I, was synthesized and studied by 2D proton NMR. The conformational properties of the peptide were characterized at different temperatures in pure aqueous solution and in a TFE/H2O (1:4 v/v) mixture. In pure aqueous solution, the peptide adopts a preferred conformation, comprising both elements of native and nonnative structures. A high alpha helix content is present in the DADELRA segment, which corresponds to an initiation site in the middle of the native alpha helix sequence. At the N-terminus flanking region, a particular nonnative folding is revealed by the J(NH-CH alpha) coupling constants and a set of unusual NOE connectivities which correspond to a helix interrupt at the first D residue. Addition of relatively small amount of TFE restores the native helix fold at the C-terminus but not at the N-terminus. On the contrary, the nonnative N-terminus structure is clearly stabilized by TFE. Our data indicate that this structure comprises (i) an Asp5-x-x-Glu8 N-terminal capping box, as recently named by Harper and Rose [Harper, E. T., & Rose, G. D. (1993) Biochemistry 32, 7605-7609], (ii) a (i,i + 3) Asp7-x-x-Arg10 salt bridge, and (iii) a hydrophobic cluster centered on Phe4 which mainly interacts with Leu9 but also with Ala2, Ala6, and Ala12 in a dynamic way. This structure is rather stable since it is still observed at 293 K in aqueous solution and 313 K in the presence of TFE. It constitutes a very potent initiation site of the alpha helix structure. This is, however, a nonnative structure involving highly conserved residues in the whole annexin family and thus may play an important role in the folding pathway as a transient "compacting helper".

Multiple interconverting conformers of the cyclic tetrapeptide tentoxin, [cyclo-(L-MeAla1-L-Leu2-MePhe[(Z) delta]3-Gly4)], as seen by two-dimensional 1H-nmr spectroscopy.

The conformations of the phytotoxic cyclic tetrapeptide tentoxin [cyclo-(L-MeAla1-L-Leu2-MePhe[(Z) delta]3-Gly4)] have been studied in aqueous solution by two-dimensional proton nmr at various temperatures. Contrary to what is observed in chloroform, tentoxin exhibits multiple exchanging conformations in water. Aggregation phenomena were also observed. Four conformations with different proportions (51, 37, 8, and 4%) were observed at -5 degrees C. Models were constructed from nmr parameters and restrained molecular dynamics simulations. All the models exhibit cis-trans-cis-trans conformation of the amide bond sequence. The conversion from one form to another is accomplished by a conformational peptide flip consisting of a 180 degree rotation of a nonmethylated peptide bond.

Glucose metabolism and internal pH of Lactococcus lactis subsp. lactis cells utilizing NMR spectroscopy.

The metabolism of glucose was studied in Lactococcus lactis subsp. CNRZ 125 by 13C NMR. The initial rate of glucose utilization was higher for exponential phase cells than for stationary phase cells [150 vs 85 nmol g (dry wt)-1 s -1]. 31P NMR was used to determine changes in glycolytic phosphorylated intermediates (fructose-1,6-diphosphate, dihydroxyacetone phosphate and phosphoglycerate). The internal pHs of L. lactis subsp. lactis CNRZ 141 and CNRZ 125 were also measured by 31P NMR as a function of the external pH during growth. When the external pH was 6.8, the internal pHs of strain CNRZ 141 and CNRZ 125 were similar, 7.4. After the external pH had decreased to 5.5, the internal pH of strain CNRZ 141 had declined by 0.6 unit, whereas that of strain CNRZ 125 had decreased by only 0.2 unit of pH.

2D 1H-NMR conformational study of phosphatidylserine diluted in perdeuterated dodecylphosphocholine micelles. Evidence for a pH-induced conformational transition.

The conformation of phosphatidylserine (DMPS) diluted in perdeuterated dodecylphosphocholine micelles (DPC) has been investigated by 1D and 2D proton NMR spectroscopy. Chemical shift pH dependence showed that the pK relative to the serine carboxyl titration (3.4 +/- 0.05) was nearly identical to that measured in bilayers. Chemical shift and NOE data revealed that the phosphatidylserine molecule undergoes a conformational transition upon titration of the serine carboxyl group. The NOE network observed between the different parts of the molecule was sufficiently abundant to allow, in combination with molecular modeling methods, an assessment of the conformational changes. The conformational changes mainly involve the glycerol backbone, which is parallel to the whole molecule, that is, to the layer normal, at low pH and becomes perpendicular to the whole molecule at neutral pH. In both cases, the conformations are remarkably close to those observed for the crystal forms of zwitterionic and negatively charged phospholipids. Two-dimensional proton NMR study of phospholipids, diluted in perdeuterated DPC micelles, appears to be a simple and relevant method to obtain complete and direct information on their conformations in a model membrane-solution interface.