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Results for “Solid-state NMR”

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At least 19 recordsLinked to original sources

PubMed1992

Melittin-induced changes in lipid multilayers. A solid-state NMR study.

Solid-state 1H, 13C, 14N, and 31P NMR spectroscopy was used to study the effects of the bee venom peptide, melittin, on aligned multilayers of dimyristoyl-, dilauryl- and ditetradecyl-phosphatidylcholines above the gel to liquid-crystalline transition temperature, Tc. Both 31P spectra from the lipid headgroups and 1H resonances from the lipid acyl chain methylene groups indicate that the peptide does not affect the mosaic spread of the lipid molecules at lipid:peptide molar ratios of 10:1, or higher. None of the samples prepared above Tc showed any evidence of the formation of hexagonal or isotropic phases. Melittin-induced changes in the chemical shift anisotropy of the headgroup phosphate and the lipid carbonyl groups, and in the choline 14N quadrupole splittings, show that the peptide has effects on the headgroup order and on the molecular organization in the sections of the acyl chains nearest to the bilayer surface. The spin-lattice relaxation time for the lipid acyl chain methylene protons was found to increase and the rotating-frame longitudinal relaxation time to markedly decrease with the addition of melittin, suggesting that motions on the nanosecond time scale are restricted, whereas the slower, collective motions are enhanced in the presence of the peptide.

Amino Acid Sequence↗

PubMed2024

Advances in solid-state NMR methods for studying RNA structures and dynamics.

Ribonucleic acid (RNA) structures and dynamics play a crucial role in elucidating RNA functions and facilitating the design of drugs targeting RNA and RNA-protein complexes. However, obtaining RNA structures using conventional biophysical techniques, such as X-ray crystallography and solution nuclear magnetic resonance (NMR), presents challenges due to the inherent flexibility and susceptibility to degradation of RNA. In recent years, solid-state NMR (SSNMR) has rapidly emerged as a promising alternative technique for characterizing RNA structure and dynamics. SSNMR has several distinct advantages, including flexibility in sample states, the ability to capture dynamic features of RNA in solid form, and suitability to character RNAs in various sizes. Recent decade witnessed the growth of 1H-detected SSNMR methods on RNA, which targeted elucidating RNA topology and base pair dynamics in solid state. They have been applied to determine the topology of RNA segment in human immunodeficiency virus (HIV) genome and the base pair dynamics of riboswitch RNA. These advancements have expanded the utility of SSNMR techniques within the RNA research field. This review provides a comprehensive discussion of recent progress in 1H-detected SSNMR investigations into RNA structure and dynamics. We focus on the established 1H-detected SSNMR methods, sample preparation protocols, and the implementation of rapid data acquisition approaches.

Dynamics↗

PubMed1992

Solid-state NMR assessment of enzyme active center structure under nonaqueous conditions.

By using solid-state NMR spectroscopy, the integrity of the active center of alpha-chymotrypsin was investigated under a variety of nonaqueous conditions. Specifically, 13C cross-polarization/magic angle spinning NMR was used to analyze the ability of alpha-chymotrypsin to stabilize a transition state intermediate analog after freezing, drying, and addition of organic solvents (both anhydrous and hydrated) to the resultant powder. Lyophilization disrupted 42 +/- 5% of the active centers; it was determined that this occurred during drying, as opposed to freezing. Seven anhydrous solvents caused 0-50% additional disruption, which occurred immediately on addition of the solvent to the enzyme powder. The extent of structural integrity loss correlated with the solvent hydrophobicity, indicating that further dehydration, i.e. stripping of water retained by the enzyme during lyophilization, was the cause. Enzyme samples prepared with lyoprotecting additives, sucrose and ammonium sulfate, exhibited varying degrees of stabilization against the drying step of lyophilization. Moreover, when hydrophilic anhydrous solvents, which had the highest propensity to strip bound water, were added to the resultant enzyme powders, no additional damage occurred.

Amino Acid Sequence↗

PubMed1992

Characterization of magnetically orientable bilayers in mixtures of dihexanoylphosphatidylcholine and dimyristoylphosphatidylcholine by solid-state NMR.

Mixtures of long-chain and short-chain phosphatidylcholine (PC) were characterized by multinuclear (13C, 31P, 2H) solid-state nuclear magnetic resonance. This work complements and extends previous characterization of such mixtures by focusing on concentrated mixtures at temperatures above the gel to liquid crystalline phase transition temperature (Tm) of the long-chain PC component. Above Tm it was observed that highly oriented, bilayer-like assemblies could be formed of mixtures of dimyristoylphosphatidylcholine (DMPC) and dihexanoylphosphatidylcholine (DHPC) in molar ratios ranging from approximately 1:3.5 to 1:2 (DHPC:DMPC) over a considerable range of lipid concentrations (at least 3-40% w/v total lipid, for a 1:2.5 sample). Orientation was observed to occur only in an L alpha-like phase. The NMR data can be accounted for by a general model of the DHPC-DMPC aggregates in which DHPC can be found in two distinct populations (one highly ordered, one not). The averaged conformations of the glycerol backbone/headgroup regions of the long- and short-chain PC composing the assemblies were judged by solid-state 13C NMR to be similar to each other. The information gleaned about these mixtures and the quality of the oriented NMR spectra obtained suggest that DHPC-DMPC mixtures may prove to be useful as model membrane media in solid-state NMR studies of biomembranes.

Dimyristoylphosphatidylcholine↗

PubMed1992

Solid-state 27Al NMR studies of aluminophosphate molecular sieves. Enhanced resolution by quadrupole nutation and double-rotation.

Solid-state 27Al NMR spectra of several aluminophosphate molecular sieves have been recorded with conventional magic-angle spinning (MAS), double-rotation (DOR) and quadrupole nutation with fast MAS. Enhanced resolution was obtained in the quadrupole nutation experiment at certain radiofrequency pulse strengths. This extra resolution can be comparable to that attainable using DOR, and does not introduce spinning sidebands.

Aluminum↗

PubMed1992

Structure determination of the cyclohexene ring of retinal in bacteriorhodopsin by solid-state deuterium NMR.

The orientation and conformation of retinal within bacteriorhodopsin of the purple membrane of Halobacterium halobium was established by solid-state deuterium NMR spectroscopy, through the determination of individual chemical bond vectors. The chromophore ([2,4,4,16,16,17,17,17,18,18-2H11]retinal) was specifically deuterium-labeled on the cyclohexene ring and incorporated into the protein. A uniaxially oriented sample of purple membrane patches was prepared and measured at a series of inclinations relative to the spectrometer field. 31P NMR was used to characterize the mosaic spread of the oriented sample, and computer simulations were applied in the analysis of the 2H NMR and 31P NMR spectral line shapes. From the deuterium quadrupole splittings, the specific orientations of the three labeled methyl groups on the cyclohexene ring could be calculated. The two adjacent methyl groups (on C1) of the retinal were found to lie approximately horizontal in the membrane and make respective angles of 94 degrees +/- 2 degrees and 75 degrees +/- 2 degrees with the membrane normal. The third group (on C5) points toward the cytoplasmic side with an angle of 46 degrees +/- 3 degrees. These intramolecular constraints indicate that the cyclohexene ring lies approximately perpendicular to the membrane surface and that it has a (6S)-trans conformation. From the estimated angle of the tilt of the chomophore long axis, it is concluded that the polyene chain is slightly curved downward to the extracellular side of the membrane.

Bacteriorhodopsins↗

PubMed1992

Solid-state 31P NMR spectroscopy of bone and bone substitutes.

Using magic angle sample spinning 31P NMR spectroscopy, we studied NMR parameters of hydroxyapatite, calcium-deficient hydroxyapatite, and beta-tricalcium phosphate. These data were compared to rabbit bone parameters. Rabbit bone spectrum is similar to deficient hydroxyapatite spectrum but high-power proton decoupling did not modify its linewidth. Moreover, both bone T1 and deficient hydroxyapatite T1 were much longer than pure hydroxyapatite T1 and beta-tricalcium phosphate T1. We used these results to develop quantitative analysis of newly deposited bone inside beta-tricalcium phosphate implants in rabbit. This method made it possible to demonstrate the effect of porosity on osteoinduction of biomaterials.

Animals↗

PubMed1992

Model ion channels: gramicidin and alamethicin.

We have discussed in some detail a variety of experimental studies which were designed to elucidate the conformational and dynamic properties of gramicidin and alamethicin. Although the behavior of these peptides is by no means fully characterized, these studies have already permitted aspects of ion channel activity to be understood in molecular terms. Studies with gramicidin in a variety of organic solutions have revealed conformational heterogeneity of this peptide; at least five major isomers exist, several of which have been characterized in detail using NMR spectroscopy and X-ray crystallography. When added to lipid membranes gramicidin undergoes a further conformational conversion. Although the conformation of gramicidin in membranes is not as well characterized as the solution conformation(s) and an X-ray structure is not yet available, detailed data, particularly from solid-state NMR studies, continue to become available and a right-handed beta 6.3 helical conformation of the peptide backbone is now generally accepted. Two of these beta 6.3 helices joined at their N-termini are believed to form the conducting channel. The conformational behavior of the side-chains of gramicidin in the membrane-bound form is not well established and several NMR, CD, fluorescence and theoretical studies are now focussed on this. Although the side-chains do not directly contact the permeating ions, they can have distinct effects on conductance and selectivity by altering the electrostatic environment sensed by the ion. The dynamics of both side-chain and backbone conformations of gramicidin appear critical to a detailed understanding of the ion transport process in this channel. As the description of the membrane-bound conformation of gramicidin becomes more detailed, simulations of ion transport using computational methods are likely to improve and will further our understanding of the processes of ion transport. As well as internal motion of the backbone and side-chains, gramicidin undergoes rotational and translational motion in the plane of the membrane. These motions do not appear to be essential for the process of ion transport but can affect channel lifetime since lifetime is determined by the rate of association and dissociation of gramicidin monomers. Gramicidin-membrane interactions are also likely to be involved in the frequency of occurrence of channel subconductance states, the frequency of channel flickering and fundamentally in the stability of the membrane-bound gramicidin conformation. Alamethicin forms channels in membranes which are strongly voltage-dependent. The molecular origin of voltage-dependent conductances has been a fundamental problem in biophysics for many years.(ABSTRACT TRUNCATED AT 400 WORDS)

Alamethicin↗

PubMed1978

Head-group conformation in phospholipids: a phosphorus-31 nuclear magnetic resonance study of oriented monodomain dipalmitoylphosphatidylcholine bilayers.

Angular-dependent 31P NMR spectra of oriented biaxial monodomain DPPC.H2O multilayers are employed to study head-group conformation in this phospholipid. The results indicate that the O-P-O plane of the phosphate, where the O's are the nonesterified oxygens of the phosphodiester, is tilted at 47 +/- 5 degrees with respect to the bilayer normal. This PO4 orientation could result in the choline moiety being extended parallel to the bilayer plane, and it will explain the breadth of the axially symmetric 31P powder spectrum observed for DPPC in excess water. This work is the first direct observation of this conformation for lecithins and it illustrates the utility of high-resolution solid-state NMR in structural studies of disordered systems.

Magnetic Resonance Spectroscopy↗

PubMed1992

A high-resolution solid-state 13C-NMR study on crystalline bovine heart cytochrome-c oxidase and lysozyme. Dynamic behavior of protein and detergent in the complex.

We have recorded 100.6-MHz high-resolution solid-state 13C-NMR spectra of crystalline cytochrome-c oxidase from bovine heart muscle and hen egg-white lysozyme, to compare conformation and dynamics of a typical membrane-protein complex with those of lysozyme. The absence of severe interference with the solid-state 13C-NMR spectra, from both the line broadenings from paramagnetic centers and overlapping of intense detergent signals, provided spectral resolution of 13C-NMR feature of cytochrome-c oxidase crystals comparable to that of lysozyme crystal and better than that of dissolved or lyophilized samples. In fact, the observed peak intensities of the polar heads of the detergents BL8SY and Brij 35 were only about 10% and 3% of the anticipated values, respectively. The dynamic behavior of the backbone and side chains of cytochrome-c oxidase was compared with that of lysozyme on the basis of the 13C spin-lattice relaxation times (T1): the backbone of the cytochrome-c oxidase turned out to be more flexible than that of lysozyme. Molecular motions of the detergent molecules attached to the proteins are found to be highly heterogeneous. Detergent molecules undergo rapid tumbling motions in the crystals in about 10 ns as detected by T1. In addition to rapid motions, slow motions were detected by 1H spin-lattice relaxation time in the rotating frame (TH1 rho) and cross-polarization time (TCH), together with data from static spectra, indicating that the aliphatic portion of the detergent interacts more strongly with hydrophobic protein surfaces than do the polar heads.

Crystallography↗

PubMed1992

Two-dimensional J-resolved and SUPERCOSY NMR experiments in the solid state.

Two 2D experiments, novel to solid-state NMR, are demonstrated using the trimethylsilyl ester of cubic octameric silicate (Q8M8). J-Resolved 13C NMR with BLEW-12 proton homonuclear decoupling reveals the scalar 13C-1H couplings and 29Si SUPERCOSY the connectivities of silicon atoms in the distorted Q8 cube.

Carbon↗

PubMed1992

Asp85 is the only internal aspartic acid that gets protonated in the M intermediate and the purple-to-blue transition of bacteriorhodopsin. A solid-state 13C CP-MAS NMR investigation.

High-resolution solid-state 13C NMR spectra of the ground state and M intermediate of the bacteriorhodopsin mutant D96N with the isotope label at [4-13C]Asp and [11-13C]Trp were recorded. The NMR spectra show that Asp85 is protonated in the M intermediate. The environment of Asp85 is quite hydrophobic. On the other hand, Asp212 remains deprotonated and a slight shift to lower field indicates a more hydrophilic environment. Asp85 also protonates in the purple-to-blue transition of bacteriorhodopsin in the deionized membrane, where it experiences a similar environment to M. The shift of Trp resonances in M reflect a conformational change of the protein in forming the M intermediate.

Aspartic Acid↗

PubMed1992

A study of tin dioxide and antimony tetroxide supported vanadium oxide catalysts by solid-state 51V and 1H NMR techniques.

A series of vanadia catalysts with various V2O5 loadings supported on SnO2 and alpha-Sb2O4 are investigated by the application of X-ray diffraction and solid-state 51V and 1H NMR techniques. XRD results show no evidence for the formation of a crystalline vanadia phase on both supports. However, the 51V NMR spectra of the catalysts reveal the existence of two types of vanadia species on the surface of the support: one due to a dispersed vanadia phase at lower vanadia loadings and the other due to a crystalline vanadia phase at higher vanadium content. The quantity of the dispersed vanadia phase, however, depends on the nature of the support material. The 1H NMR results provide evidence for the existence or non-existence of a metal oxide support interaction through the support surface hydroxyl groups.

Antimony↗

PubMed1992

Orientation of the valine-1 side chain of the gramicidin transmembrane channel and implications for channel functioning. A 2H NMR study.

The orientation of the valine-1 side chain of gramicidin was determined by solid-state 2H NMR using valine-1-deuterated (d8) gramicidin. The peptide was incorporated into DMPC bilayers that were oriented between glass plates. When the plates were oriented with their normal perpendicular to the magnetic field, four quadrupolar splittings were observed of 106, 68, 9.7, and 2.0 kHz. These resonances were assigned to C alpha D, C beta D, and the deuterons of each of the C gamma D3 methyl groups, respectively. The average orientation of the various C-D bonds was calculated with respect to the helix axis. The angle obtained for the C alpha-D resonance was consistent with a single-stranded beta 6.3-helical model for the backbone but not with double-helical models. The angles of the side chain were then fitted to a model for the right-handed beta 6.3-helix. Rotation of the valine-1 side chain yielded a set of torsion angles that matched the angles as determined from the 2H NMR measurements. The corresponding orientation of the valine-1 side chain (chi 1 = -5 degrees) was found to be quite unusual, but it explains well the importance of a branched side chain at position 1 for channel formation and stability. A van der Waals interaction between valine-1 of one monomer and alanine-5 of the other helps to stabilize the gramicidin dimer.

Amino Acid Sequence↗

PubMed1992

Interfacial conformation of dipalmitoylglycerol and dipalmitoylphosphatidylcholine in phospholipid bilayers.

Diacylglycerols are minor constituents of membrane lipids, yet are essential in the activation and membrane association of protein kinase C. Solid-state 13C NMR experiments have been used to characterize the orientation of the glycerol backbone of dipalmitoylglycerol (DPG) and dipalmitoylphosphatidylcholine (DPPC) in egg phosphatidylcholine (PC) bilayers. The 13C NMR spectra of both DPG and DPPC specifically 13C-labeled at the sn-2 chain carbonyl exhibit a single narrow resonance (approximately 2 ppm) in liquid-crystalline egg PC bilayers. In contrast, specific 13C-labeling of both the sn-1 and sn-2 chain carbonyls results in an additional broad component (24-32 ppm) with an axially symmetric line shape. These data reveal that DPG has a distinct motionally-averaged structure in PC bilayers that is similar to that of DPPC and is not significantly affected by the absence of the large polar PC headgroup. The NMR line shapes are roughly consistent with the results of previous FTIR and NMR studies that indicate the sn-1 chain extends from the C1 carbon of the glycerol backbone into the hydrophobic interior of the bilayer, while the sn-2 chain first extends parallel to the bilayer surface and incorporates a bend at the ester linkage in order to keep the sn-1 and sn-2 chains parallel. However, the data suggest that the time-averaged orientation of the glycerol backbone is tilted from the bilayer normal, in contrast to the nearly parallel orientation observed in the crystal structures of phosphatidylcholines and phosphatidylethanolamines or the perpendicular orientation observed in the crystal structures of diacylglycerols.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1992

Synthesis of backbone deuterium labelled [r(CGCGAAUUCGCG)]2 and HPLC purification of synthetic RNA.

The chemical synthesis of backbone deuterium labelled [r(CGCGAAU*U*CGCG)]2 (U* = [5'-2H]U) is described. An efficient purification procedure was developed using a polymeric reverse phase (PRP) HPLC column at 60 degrees C. This procedure provided pure RNA dodecamer in the multi-milligram quantities (39% overall yield) necessary for dynamics studies using solid-state deuterium NMR. The purification method has been effectively applied to other RNA sequences and will assist biophysical studies which require relatively large quantities of RNA oligomers.

Amides↗

PubMed1992

Conformation-dependent change in antitumor activity of linear and branched (1----3)-beta-D-glucans on the basis of conformational elucidation by carbon-13 nuclear magnetic resonance spectroscopy.

The antitumor activity of (1----3)-beta-D-glucans was tested in order to clarify its conformation-dependent response together with conformational elucidation by carbon-13 nuclear magnetic resonance (13C-NMR) spectroscopy. It was shown that the following three conformations, single chain, single helix and triple helix, are readily distinguished by the high-resolution solid-state 13C-NMR method. It turned out that preparations of linear (1----3)-beta-D-glucans of a triple helical conformation were ineffective in the inhibition of tumor growth. These linear (1----3)-beta-D-glucans were converted to an effective form in the inhibition of tumor growth when they were lyophilized from dimethyl sulfoxide (DMSO) solutions as a result of a conformational change from the triple helical to the single chain forms. They were not effective, however, when assayed in DMSO solution. In contrast, it was found that a branched (1----3)-beta-D-glucan is effective not only in either saline solutions of the triple helical sample or the lyophilized sample from DMSO, but also in DMSO solution. The aforementioned drastic change in antitumor activity was interpreted in terms of resulting conformational changes as analyzed by the 13C-NMR method.

Animals↗

PubMed1992

13C magic angle spinning NMR study of the light-induced and temperature-dependent changes in Rhodobacter sphaeroides R26 reaction centers enriched in [4'-13C]tyrosine.

Solid-state 13C magic angle spinning (MAS) NMR has been used to investigate detergent-solubilized photosynthetic reaction centers of Rhodobacter sphaeroides R26, selectively enriched in [4-13C]-tyrosine. The reaction centers were frozen, in the dark and while subject to intense illumination, and studied at temperatures between approximately 215 and approximately 260 K. The signal consists of at least seven narrow lines superimposed on a broad doublet. The chemical shift anisotropy is similar to that for crystalline tyrosine. The two narrowest resonances, corresponding to signals from individual tyrosines, are 28 +/- 5 Hz wide, comparable to what is observed for quaternary carbons in linearly elastic organic solids. The line width as well as the chemical shift of these signals is essentially independent of temperature. This provides strong evidence for an unusually ordered, well-shielded, and structurally, electrostatically, and thermodynamically stable interior of the protein complex without structural heterogeneities. As the temperature is lowered, additional signal from the labels develops and the natural abundance resonances from the detergent broaden, providing evidence for considerable flexibility at the exterior of the protein complex and in the detergent belt at the higher temperatures. In addition, the NMR provides evidence for an electrostatically uniform and neutral complex, since the total dispersion in isotropic shifts for the labels is < 5 ppm and corresponds to electron density variations of less than 0.03 electronic equivalents with respect to tyrosine in the solid state or in solution. When the sample is frozen while subject to intense illumination, a substantial part of the protein is brought into the charge-separated state P.+QA.-. At least three sharp resonances, including the narrowest lines, are substantially reduced in intensity. It is argued that this effect is caused by the electronic spin density associated with the oxidized primary donor P.+. These results strongly suggest that the environment of the special pair is extremely rigid and question the role of protein conformational distortions during the primary photoprocess.

Carbon Isotopes↗