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1.
Infrared reflection-absorption spectroscopy (IRRAS) of lipid/protein monolayer films in situ at the air/water interface provides unique molecular structure and orientation information from the film constituents. The technique is thus well suited for studies of lipid/protein interaction in a physiologically relevant environment. Initially, the nature of the IRRAS experiment is described and the molecular structure information that may be obtained is recapitulated. Subsequently, several types of applications, including the determination of lipid chain conformation and tilt as well as elucidation of protein secondary structure are reviewed. The current article attempts to provide the reader with an understanding of the current capabilities of IRRAS instrumentation and the type of results that have been achieved to date from IRRAS studies of lipids, proteins, and lipid/protein films of progressively increasing complexity. Finally, possible extensions of the technology are briefly considered. 相似文献
2.
Solid-state NMR as a method to reveal structure and membrane-interaction of amyloidogenic proteins and peptides 总被引:1,自引:0,他引:1
It is important to understand the Amyloid fibril formation in view of numerous medical and biochemical aspects. Structural determination of amyloid fibril has been extensively studied using electron microscopy. Subsequently, solid state NMR spectroscopy has been realized to be the most important means to determine not only microscopic molecular structure but also macroscopic molecular packing. Molecular structure of amyloid fibril was first predicted to be parallel beta-sheet structure, and subsequently, was further refined for Abeta(1-40) to be cross beta-sheet with double layered in register parallel beta-sheet structure by using solid state NMR spectroscopy. On the other hand, anti-parallel beta-sheet structure has been reported to short fragments of Abeta-amyloid and other amyloid forming peptides. Kinetic study of amyloid fibril formation has been studied using a variety of methods, and two-step autocatalytic reaction mechanism used to explain fibril formation. Recently, stable intermediates or proto-fibrils have been observed by electron microscope (EM) images. Some of the intermediates have the same microscopic structure as the matured fibril and subsequently change to matured fibrils. Another important study on amyloid fibril formation is determination of the interaction with lipid membranes, since amyloid peptide are cleaved from amyloid precursor proteins in the membrane interface, and it is reported that amyloid lipid interaction is related to the cytotoxicity. Finally it is discussed how amyloid fibril formation can be inhibited. Firstly, properly designed compounds are reported to have inhibition ability of amyloid fibril formation by interacting with amyloid peptide. Secondly, it is revealed that site directed mutation can inhibit amyloid fibril formation. These inhibitors were developed by knowing the fibril structure determined by solid state NMR. 相似文献
3.
It is important to understand the Amyloid fibril formation in view of numerous medical and biochemical aspects. Structural determination of amyloid fibril has been extensively studied using electron microscopy. Subsequently, solid state NMR spectroscopy has been realized to be the most important means to determine not only microscopic molecular structure but also macroscopic molecular packing. Molecular structure of amyloid fibril was first predicted to be parallel β-sheet structure, and subsequently, was further refined for Aβ(1-40) to be cross β-sheet with double layered in register parallel β-sheet structure by using solid state NMR spectroscopy. On the other hand, anti-parallel β-sheet structure has been reported to short fragments of Aβ-amyloid and other amyloid forming peptides. Kinetic study of amyloid fibril formation has been studied using a variety of methods, and two-step autocatalytic reaction mechanism used to explain fibril formation. Recently, stable intermediates or proto-fibrils have been observed by electron microscope (EM) images. Some of the intermediates have the same microscopic structure as the matured fibril and subsequently change to matured fibrils. Another important study on amyloid fibril formation is determination of the interaction with lipid membranes, since amyloid peptide are cleaved from amyloid precursor proteins in the membrane interface, and it is reported that amyloid lipid interaction is related to the cytotoxicity. Finally it is discussed how amyloid fibril formation can be inhibited. Firstly, properly designed compounds are reported to have inhibition ability of amyloid fibril formation by interacting with amyloid peptide. Secondly, it is revealed that site directed mutation can inhibit amyloid fibril formation. These inhibitors were developed by knowing the fibril structure determined by solid state NMR. 相似文献
4.
During the past year, electron crystallography of membrane proteins has provided structural insights into the mechanism of several different transporters and into their interactions with lipid molecules within the bilayer. From a technical perspective there have been important advances in high-throughput screening of crystallization trials and in automated imaging of membrane crystals with the electron microscope. There have also been key developments in software, and in molecular replacement and phase extension methods designed to facilitate the process of structure determination. 相似文献
5.
Phospholipid component volumes: determination and application to bilayer structure calculations. 总被引:4,自引:2,他引:2
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We present a new method for the determination of bilayer structure based on a combination of computational studies and laboratory experiments. From molecular dynamics simulations, the volumes of submolecular fragments of saturated and unsaturated phosphatidylcholines in the liquid crystalline state have been extracted with a precision not available experimentally. Constancy of component volumes, both among different lipids and as a function of membrane position for a given lipid, have been examined. The component volumes were then incorporated into the liquid crystallographic method described by Wiener and White (1992. Biophys. J. 61:434-447, and references therein) for determining the structure of a fluid-phase dioleoylphosphatidylcholine bilayer from x-ray and neutron diffraction experiments. 相似文献
6.
M Stephen Trent 《Biochimie et biologie cellulaire》2004,82(1):71-86
Lipopolysaccharide (LPS) is the major surface molecule of Gram-negative bacteria and consists of three distinct structural domains: O-antigen, core, and lipid A. The lipid A (endotoxin) domain of LPS is a unique, glucosamine-based phospholipid that serves as the hydrophobic anchor of LPS and is the bioactive component of the molecule that is associated with Gram-negative septic shock. The structural genes encoding the enzymes required for the biosynthesis of Escherchia coli lipid A have been identified and characterized. Lipid A is often viewed as a constitutively synthesized structural molecule. However, determination of the exact chemical structures of lipid A from diverse Gram-negative bacteria shows that the molecule can be further modified in response to environmental stimuli. These modifications have been implicated in virulence of pathogenic Gram-negative bacteria and represent one of the molecular mechanisms of microbial surface remodeling used by bacteria to help evade the innate immune response. The intent of this review is to discuss the enzymatic machinery involved in the biosynthesis of lipid A, transport of the molecule, and finally, those enzymes involved in the modification of its structure in response to environmental stimuli. 相似文献
7.
Aurélien Lorin Danny Létourneau Andrée Lefebvre Jean-Guy LeHoux Pierre Lavigne 《Biomolecular NMR assignments》2013,7(1):21-24
Steroidogenic acute regulatory (StAR)—related lipid transfer proteins possess a START (steroidogenic acute regulatory-related lipid transfer) domain. START domains are conserved protein modules involved in the non-vesicular intracellular transport of lipids and cholesterol in mammals. Fifteen mammalian proteins, divided in five subfamilies, are reported to possess a START domain. Members of the STARD4 subfamily, i.e. STARD4, 5 and 6 are essentially single START domains and are thought to be involved in the intracellular transport of cholesterol. No structure of a cholesterol-bound START domain from this family has been resolved yet. The determination of the structure of such a complex would contribute to a better understanding of the mechanism of ligand binding and transport by START domains, two unresolved aspects of their structural biology. In this context, we have undertaken the structure determination of a ligand-bound form of STARD5 by NMR. Here, we report the 1H, 13C and 15N backbone resonance assignments of the ligand-free STARD5. 相似文献
8.
L V Antonenko Iu D Kholodova L I Apukhovskaia P A Voziian E V Solodova L V Kvashnina 《Ukrainski? biokhimicheski? zhurnal》1990,62(3):54-59
The lipid composition and structure of low- and high-density lipoproteins (LDL and HDL) have been studied in children with the vitamin D-deficient rachitis. An increase of the cholesterol content in the atherogenic LDL against a background of its decrease in the antiatherogenic HDL has been revealed, that is analogous to the observed changes under the atherosclerosis development in adults. Due to the studies in the amino acidic composition and determination of the nature and quantity of the charged groups of lipoprotein particles accessible for titration definite, certain disturbance in the structure of lipoproteins are observed as such that may cause disturbances in their functional activity. 相似文献
9.
Apolipoprotein H (ApoH), also known as beta(2)-glycoprotein I, is a plasma glycoprotein with its in vivo physiological and pathogenic roles being closely related to its interaction with negatively charged membranes. Although the three-dimensional crystal structure of ApoH has been recently solved, direct evidence about the spatial state of ApoH on the membrane is still lacking. In this work, the interactions of ApoH with the lipid layer are studied by a combination of lipid monolayer approach and surface concentration determination. The spatial state of the orientation of ApoH on the lipid layer is investigated by analyzing the process of membrane-attached ApoH molecules being extruded out from the phospholipid monolayer by compression. The results show that on neutral lipid layer ApoH has an upright orientation, which is not sensitive to the phase state of the lipid layer. However, on acidic lipid layer, ApoH may have two forms of orientation. One is an upright orientation in the liquid phase region, and the other is flat orientation on the condensed domain region. The variation of the spatial state of ApoH on the lipid layer may relate to a variety of its physiological functions. 相似文献
10.
Anna Diller Cécile Loudet Fabien Aussenac Gérard Raffard Sylvie Fournier Michel Laguerre Axelle Grélard Stanley J. Opella Francesca M. Marassi Erick J. Dufourc 《Biochimie》2009
Major biological processes occur at the biological membrane. One of the great challenges is to understand the function of chemical or biological molecules inside the membrane; as well of those involved in membrane trafficking. This requires obtaining a complete picture of the in situ structure and dynamics as well as the topology and orientation of these molecules in the membrane lipid bilayer. These led to the creation of several innovative models of biological membranes in order to investigate the structure and dynamics of amphiphilic molecules, as well as integral membrane proteins having single or multiple transmembrane segments. Because the determination of the structure, dynamics and topology of molecules in membranes requires a macroscopic alignment of the system, a new membrane model called ‘bicelles’ that represents a crossover between lipid vesicles and classical micelles has become very popular due to its property of spontaneous self-orientation in magnetic fields. In addition, crucial factors involved in mimicking natural membranes, such as sample hydration, pH and salinity limits, are easy to control in bicelle systems. Bicelles are composed of mixtures of long chain (14–18 carbons) and short chain phospholipids (6–8 carbons) hydrated up to 98% with buffers and may adopt various morphologies depending on lipid composition, temperature and hydration. We have been developing bicelle systems under the form of nano-discs made of lipids with saturated or biphenyl-containing fatty acyl chains. Depending on the lipid nature, these membranous nano-discs may be macroscopically oriented with their normal perpendicular or parallel to the magnetic field, providing a natural ‘molecular goniometer’ for structural and topological studies, especially in the field of NMR. Bicelles can also be spun at the magic angle and lead to the 3D structural determination of molecules in membranes. 相似文献
11.
Fluorescence quenching and resonance energy transfer methods have been used to investigate the position of fluorophores in the lateral and transverse planes of the lipid bilayer. A series of n-(9-anthroyloxy) fatty acids (n = 2, 6, 9, and 12) have been used as energy-transfer acceptors so that apparent transfer distances from a membrane-bound donor (N-stearoyltryptophan) have a transverse as well as a lateral component. Both theory and experiment show that the energy-transfer method is not precise enough to discriminate between the positions of the fluorophores in the transverse plane of the bilayer. The n-(9-anthroyloxy) fatty acids are also susceptible to quenching by the indole moiety of tryptophan. The relative quenching efficiency can provide a semiquantitative measure of the position of quenching molecules in the lipid bilayer. The quenching techniques are applied to the determination of the orientation of gramicidin A in lipid bilayers. The tryptophan residues of gramicidin appear to be located near the membrane surface in agreement with the head-to-head dimeric structure proposed by D. W. Urry et al. [(1971) Proc. Natl. Acad. Sci. U.S.A. 68, 672--676]. 相似文献
12.
A simple approach to membrane protein secondary structure and topology based on NMR spectroscopy
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Marassi FM 《Biophysical journal》2001,80(2):994-1003
This paper describes a simple, qualitative approach for the determination of membrane protein secondary structure and topology in lipid bilayer membranes. The approach is based on the observation of wheel-like resonance patterns observed in the NMR 1H-15N/15N polarization inversion with spin exchange at the magic angle (PISEMA) and 1H/15N heteronuclear correlation (HETCOR) spectra of membrane proteins in oriented lipid bilayers. These patterns, named Pisa wheels, have been previously shown to reflect helical wheel projections of residues that are characteristic of alpha-helices associated with membranes. This study extends the analysis of these patterns to beta-strands associated with membranes and demonstrates that, as for the case of alpha-helices, Pisa wheels are extremely sensitive to the tilt, rotation, and twist of beta-strands in the membrane. Therefore, the Pisa wheels provide a sensitive, visually accessible, qualitative index of membrane protein secondary structure and topology. 相似文献
13.
We present a methodology of lipid nanotubes (LNT) and nanodisks technologies optimized in our laboratory for structural studies of membrane‐associated proteins at close to physiological conditions. The application of these lipid nanotechnologies for structure determination by cryo‐electron microscopy (cryo‐EM) is fundamental for understanding and modulating their function. The LNTs in our studies are single bilayer galactosylceramide based nanotubes of ~20 nm inner diameter and a few microns in length, that self‐assemble in aqueous solutions. The lipid nanodisks (NDs) are self‐assembled discoid lipid bilayers of ~10 nm diameter, which are stabilized in aqueous solutions by a belt of amphipathic helical scaffold proteins. By combining LNT and ND technologies, we can examine structurally how the membrane curvature and lipid composition modulates the function of the membrane‐associated proteins. As proof of principle, we have engineered these lipid nanotechnologies to mimic the activated platelet's phosphtaidylserine rich membrane and have successfully assembled functional membrane‐bound coagulation factor VIII in vitro for structure determination by cryo‐EM. The macromolecular organization of the proteins bound to ND and LNT are further defined by fitting the known atomic structures within the calculated three‐dimensional maps. The combination of LNT and ND technologies offers a means to control the design and assembly of a wide range of functional membrane‐associated proteins and complexes for structural studies by cryo‐EM. The presented results confirm the suitability of the developed methodology for studying the functional structure of membrane‐associated proteins, such as the coagulation factors, at a close to physiological environment. Proteins 2014; 82:2902–2909. © 2014 Wiley Periodicals, Inc. 相似文献
14.
Modeling of the structural features of integral-membrane proteins reverse-environment prediction of integral membrane protein structure (REPIMPS) 总被引:1,自引:0,他引:1
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Dastmalchi S Morris MB Church WB 《Protein science : a publication of the Protein Society》2001,10(8):1529-1538
The Profiles-3D application, an inverse-folding methodology appropriate for water-soluble proteins, has been modified to allow the determination of structural properties of integral-membrane proteins (IMPs) and for testing the validity of solved and model structures of IMPs. The modification, known as reverse-environment prediction of integral membrane protein structure (REPIMPS), takes into account the fact that exposed areas of side chains for many residues in IMPs are in contact with lipid and not the aqueous phase. This (1) allows lipid-exposed residues to be classified into the correct physicochemical environment class, (2) significantly improves compatibility scores for IMPs whose structures have been solved, and (3) reduces the possibility of rejecting a three-dimensional structure for an IMP because the presence of lipid was not included. Validation tests of REPIMPS showed that it (1) can locate the transmembrane domain of IMPs with single transmembrane helices more frequently than a range of other methodologies, (2) can rotationally orient transmembrane helices with respect to the lipid environment and surrounding helices in IMPs with multiple transmembrane helices, and (3) has the potential to accurately locate transmembrane domains in IMPs with multiple transmembrane helices. We conclude that correcting for the presence of the lipid environment surrounding the transmembrane segments of IMPs is an essential step for reasonable modeling and verification of the three-dimensional structures of these proteins. 相似文献
15.
Deanna DiDonato Dawn L Brasaemle 《The journal of histochemistry and cytochemistry》2003,51(6):773-780
The study of proteins associated with lipid droplets in adipocytes and many other cells is a rapidly developing area of inquiry. Although lipid droplets are easily visible by light microscopy, few standardized microscopy methods have been developed. Several methods of chemical fixation have recently been used to preserve cell structure before visualization of lipid droplets by light microscopy. We tested the most commonly used methods to compare the effects of the fixatives on cellular lipid content and lipid droplet structure. Cold methanol fixation has traditionally been used before visualization of cytoskeletal elements. We found this method unacceptable for study of lipid droplets because it extracted the majority of cellular phospholipids and promoted fusion of lipid droplets. Cold acetone fixation is similarly unacceptable because the total cellular lipids are extracted, causing collapse of the shell of lipid droplet-associated proteins. Fixation of cells with paraformaldehyde is the method of choice, because the cells retain their lipid content and lipid droplet structure is unaffected. As more lipid droplet-associated proteins are discovered and studied, it is critical to use appropriate methods to avoid studying artifacts. 相似文献
16.
V A Roginski? 《Molekuliarnaia biologiia》1990,24(6):1582-1589
The structure, thermodynamic, and kinetic features of polyunsaturated fatty acids derivatives as the main substratum of lipid peroxidation (POL) have been considered. The heats of key POL reactions have been estimated. Kinetic consequences of these estimations have been analyzed. The dependence of POL rate on O2 concentration have been considered in detail both in the absence and in the presence of antioxidants. The essential features of POL processes in lipid bilayers resulting from the specific structure and molecular dynamics have been discussed. 相似文献
17.
18.
Lala AK 《Chemistry and physics of lipids》2002,116(1-2):177-188
This report summarizes our efforts towards depth-dependent analysis of membranes by design of suitable fluorescent and photoactivable lipid probes, which can be incorporated into membranes. The objective of depth-dependent analysis has been two fold, one to obtain information on lipid domains and other on transmembrane domains of membrane-bound proteins. In view of increasing importance of lipid rafts and other localized domain and limited success in case of structure determination of membrane-bound proteins vis-à-vis their soluble counterparts, it is tempting to rapidly attach fluorescent or photoactivable probes to lipids to get a probes where relatively little attention is paid to design of such probes. We have shown here how careful design of such probes is required to immobilize such probes in membranes for effective depth-dependent analysis of membranes. An effective design has become important when identification of putative transmembrane domains predicted primarily from the genome data based on hydropathy plots, often needs confirmation by contemporary methodology. 相似文献
19.
Structure of lipid bilayers 总被引:8,自引:0,他引:8
The quantitative experimental uncertainty in the structure of fully hydrated, biologically relevant, fluid (L(alpha)) phase lipid bilayers has been too large to provide a firm base for applications or for comparison with simulations. Many structural methods are reviewed including modern liquid crystallography of lipid bilayers that deals with the fully developed undulation fluctuations that occur in the L(alpha) phase. These fluctuations degrade the higher order diffraction data in a way that, if unrecognized, leads to erroneous conclusions regarding bilayer structure. Diffraction measurements at high instrumental resolution provide a measure of these fluctuations. In addition to providing better structural determination, this opens a new window on interactions between bilayers, so the experimental determination of interbilayer interaction parameters is reviewed briefly. We introduce a new structural correction based on fluctuations that has not been included in any previous studies. Updated measurements, such as for the area compressibility modulus, are used to provide adjustments to many of the literature values of structural quantities. Since the gel (L(beta)') phase is valuable as a stepping stone for obtaining fluid phase results, a brief review is given of the lower temperature phases. The uncertainty in structural results for lipid bilayers is being reduced and best current values are provided for bilayers of five lipids. 相似文献
20.
Direct determination of crystallographic phases for diffraction data from phospholipid multilamellar arrays. 总被引:2,自引:2,他引:0
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D L Dorset 《Biophysical journal》1990,58(5):1077-1087
Direct determination of crystallographic phases based on probabilistic of sigma 1 and sigma 2 "triplet" structure invariants has been found to be an effective technique for structure analysis with lamellar x-ray or electron diffraction intensity data from phospholipids. In many cases, nearly all phase values are determined, permitting a structure density (electron density for x-ray diffraction; electrostatic potential for electron diffraction) map to be calculated, which is directly interpretable in terms of known bilayer lipid structure. The major source of error is found to be due to the distortion of observed electron diffraction intensity data by incoherent multiple scattering, which can significantly affect the appearance of the electrostatic potential map, but not the success of the phase determination, as long as the observed Patterson function can be interpreted. 相似文献