13C cross‐polarization magic angle spinning NMR of ferrocene derivatives

The 13C chemical shifts of the CP/MAS NMR for ferrocene derivatives have been measured. Copyright © 2002 John Wiley & Sons, Ltd.     

13C/15N distance determination by CPMAS NMR in uniformly 13C labeled molecules

The REDOR and CPMAS techniques are applied for measuring 13C-15N dipolar coupling constants in glycine. It is shown that the selective CP or SPECIFIC CP technique removes the coherent evolution of the spin system under homonuclear 13C-13C J couplings. While the large coupling constant (approximately 900 Hz) is readily determined because of the presence of large oscillations in the CPMAS dynamics, their absence precludes the measurement of the small coupling constant (approximately 200 Hz). The experimental results and numerical simulations demonstrate that the determination of 13C-15N coupling constants of medium size (<1 kHz) by the CPMAS technique is mainly limited by the strength of the 1H decoupling field and the size of the 13C and 15N chemical shift anisotropies.     

Solid-phase synthesis and 1H and 13C high-resolution magic angle spinning NMR of 13C-labeled resin-bound saccharides

We show how high-resolution magic angle-spinning NMR spectroscopy can be used to characterize 13C-labeled saccharides that have been prepared using solid-phase synthesis techniques while they are still bound to a solid-support resin. With the use of 13C-labeled glucose as the starting material, we have successfully synthesized mono-, di- and trisaccharides with uniform 13C labeling of the saccharide rings. Using these materials, we have been able to assign the 13C and 1H spectra and to characterize various impurities on the resin beads.     

Heating caused by radiofrequency irradiation and sample rotation in 13C magic angle spinning NMR studies of lipid membranes

Application of rapid sample rotation and radiofrequency irradiation in magic angle spinning (MAS) NMR of lipid bilayers can significantly increase the sample temperature. In this work, we studied the extent of heating during the acquisition of 1H-decoupled 13C MAS spectra of hydrated dimyristoylphosphatidylcholine (DMPC) in the L(alpha) phase. First, we describe a simple procedure for determining the increase in temperature by observing the shift of the 1H water signal. The method is then used to identify and assess the various factors that contribute to the sample heating. The important factors discussed in this paper include: (i) the spinning speed, (ii) the variable-temperature gas pressure, (iii) the rotor geometry, (iv) the power, duration and frequency of the radiofrequency irradiation and (v) the hydration level. A comparison of different heteronuclear decoupling schemes in terms of their ability to produce highly resolved 13C spectra of DMPC is also reported.     

13C CP/MAS NMR studies of vitamin E model compounds

13C cross-polarization magic angle spinning (CP/MAS) NMR data for 2,2,5,7,8-pentamethylchroman-6-ol (2), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox c) (3) and its acetate (4), 2-methoxy-2,2,5,7,8-pentamethylchroman-6-ol (5), 2-hydroxy-2,2,5,7,8-pentamethylchroman-6-ol (6) and 2,2,5,7,8-pentamethylchroman (7) are reported. A deshielding of 7.7 ppm for the carboxylic carbon was observed in solid Trolox due to formation of intermolecular hydrogen bonds within cyclic dimers. Such crystal packing permits effective cross-polarization and fast relaxation (short T1rho(H)). The impact of the proton concentration on the CP dynamics is reflected by the longer T(CP) and T1rhoH for Trolox-d2 (deuterated at mobile proton sites). The calculated GIAO RHF shielding constants are sensitive to intramolecular effects: rotation around the C-6-O bond (changes of sigma up to 8 ppm) and conformation at C-2.     

Reaction of o-phenylenediamine with diacetyl monoxime: characterisation of the product by solid-state 13C and 15N MAS NMR

2,3-dimethylquinoxaline (DMQ) and dimethylglyoxime (DMGH2) form a 1:1 hydrogen-bonded complex in the solid state, which is completely dissociated in methanol solution. There are small differences in solid-state 13C shifts between the separated components DMQ and DMGH2 and the complex. The changes in 15N solid-state chemical shifts are more significant: the hydrogen bond imparting a low frequency shift of ca 19 ppm. The effect of direct protonation on the DMQ solid-state 15N shifts was measured, and the experimental 15N data correlated with those from GIAO molecular orbital (MO) calculations.     

High-resolution solid-state MAS 73Ge NMR spectra of some organogermanes

High-resolution solid-state magic angle spinning (73)Ge NMR spectra of some organogermanium compounds were measured. Most tetrasubstituted germanes with identical substituents exhibited signals except for one case. Tetrasubstituted germanes with two kinds of different but somewhat similar substituents exhibited broad peaks. Trisubstituted germanes failed to show signals, indicating the importance of symmetry around germanium.     

Structural characterization and quantitation of compound loading of disubstituted benzoates bound to Wang resin through high-resolution magic angle spinning NMR spectroscopy

The combination of 1D and 2D high-resolution magic angle spinning NMR experiments led to the assignment of the proton and carbon resonances for several disubstituted benzoates bound to a polystyrene resin through a Wang linker. It is shown that the signal corresponding to the methylene protons of the linker can be utilized to monitor the solid-phase reactions and determine the loading of the compounds on the resin.     

7Li and 13C solid-state NMR spectra of lithium cuprates

7Li and 13C solid-state MAS NMR spectra of three lithium cuprates with known X-ray structures--lithium([12]crown-4)2 dimethyl and diphenyl cuprate (1,2) and lithium(thf)4-[tris(trimethylsilyl) methyl]2 cuprate (3)--have been measured and analysed with respect to the quadrupolar coupling constants of lithium-7, chi(7Li), and the asymmetry parameters of the quadrupolar interactions, eta(7Li), as well as the 6, 7Li and 13C chemical shifts. The chi(7Li) values of 23, 30, and 18 kHz for 1, 2 and 3, respectively, are in line with the high symmetry around the lithium nucleus in the solvent-separated structures and may be used as reference data for this structural motif. Calculations based on charges derived from ab initio 6-31 G* HF computations using the point charge model (PCM) and the program GAMESS support the experimental findings.     

Phosphorylated amino acids: model compounds for solid-state 31P NMR spectroscopic studies of proteins

Solid-state 31P NMR spectroscopy was applied to measure the isotropic chemical shifts, chemical shift anisotropies and asymmetry parameters of three phosphorylated amino acids, O-phospho-L-serine, O-phospho-L-threonine and O-phospho-L-tyrosine. The cross-polarization buildup rates and longitudinal relaxation times of 31P and 1H were-determined and compared with the values measured for a triphosphate (GppCH2p) bound to a crystalline protein (Ras). It is shown that the phosphorylated amino acids are well-suited model compounds, e.g. for the optimization of experiments on crystalline proteins. Two-dimensional exchange experiments on O-phospho-L-tyrosine indicate the existence of an exchange between the two different conformations of the molecule.     

Assignment of 13C resonances of a nematic liquid crystal using off-magic angle spinning

A novel method for assigning the resonances in the 13C NMR spectrum of a static liquid crystalline sample in its nematic phase is proposed. The method is based on the fact that the carbon chemical shifts in the isotropic phase and in the oriented phase under static and off-magic angle spinning (OMAS) conditions are uniquely related by the tensorial property of the CSA tensor, requiring just one OMAS spectrum and the assignment in the isotropic phase. A computational procedure is proposed to take into account deviations arising out of non-ideal experimental conditions and the assignments are made by identifying the minimum in the differences in the frequencies between calculated and experimental line positions. Practical implementation of the method has also been demonstrated in the case of the liquid crystal N-(4-ethoxybenzylidene)-4-n-butylaniline.     

De Novo 3D Structure Determination from Sub‐milligram Protein Samples by Solid‐State 100 kHz MAS NMR Spectroscopy

Solid‐state NMR spectroscopy is an emerging tool for structural studies of crystalline, membrane‐associated, sedimented, and fibrillar proteins. A major limitation for many studies is still the large amount of sample needed for the experiments, typically several isotopically labeled samples of 10–20 mg each. Here we show that a new NMR probe, pushing magic‐angle sample rotation to frequencies around 100 kHz, makes it possible to narrow the proton resonance lines sufficiently to provide the necessary sensitivity and spectral resolution for efficient and sensitive proton detection. Using restraints from such spectra, a well‐defined de novo structure of the model protein ubiquitin was obtained from two samples of roughly 500 μg protein each. This proof of principle opens new avenues for structural studies of proteins available in microgram, or tens of nanomoles, quantities that are, for example, typically achieved for eukaryotic membrane proteins by in‐cell or cell‐free expression.     

Dynamic and magnetic susceptibility effects on the MAS NMR linewidth of a tetrapeptide bound to different resins

Under magic angle spinning, the NMR spectrum of the tetrapeptide Ala‐Ile‐Gly‐Met bound to a Wang resin, and swollen in DMF, exhibits proton and carbon linewidths that are sharp enough to allow the complete characterization of the peptide using classical liquid‐state NMR methods. The proton linewidths of the bound peptide remain, however, about three times larger than those of the free peptide in solution. The residual NMR linewidth originates essentially from incompletely averaged magnetic susceptibility effects due to the Wang resin. Replacing the aromatic Wang resin with a PEGA or POEPOP resin removes this effect. To investigate the contribution to line broadening of the peptide dynamics, relaxation studies were performed on the peptide bound to Wang and POEPOP resins. Copyright © 2001 John Wiley & Sons, Ltd.     

Conformation analysis and molecular mobility of ethylene and tetrafluoroethylene copolymer using solid-state 19F MAS and 1H --> 19F CP/MAS NMR spectroscopy

The changes in the conformation and molecular mobility accompanied by a phase transition in the crystalline domain were analyzed for ethylene (E) and tetrafluoroethylene (TFE) copolymer, ETFE, using variable-temperature (VT) solid-state 19F magic angle spinning (MAS) and 1H --> 19F cross-polarization (CP)/MAS NMR spectroscopy. The shifts of the signals for fluorines in TFE units to higher frequency and the continuing decrease and increase in the T1rho(F) values suggest that conformational exchange motions exist in the crystalline domain between 42 and 145 degrees C. Quantum chemical calculations of magnetic shielding constants showed that the high-frequency shift of TFE units should be induced by trans to gauche conformational changes at the CH2-CF2 linkage in the E-TFE unit. Although the 19F signals of the crystalline domain are substantially overlapped with those of the amorphous domain at ambient probe temperature (68 degrees C), they were successfully distinguished by using the dipolar filter and spin-lock pulse sequences at 145 degrees C. The dipolar coupling constants for the crystalline domain, which can be estimated by fitting the dipolar oscillation behaviors in the 1H --> 19F CP curve, showed a significant decrease with increasing temperature from 42 to 145 degrees C. This is due to the averaging of 1H-19F dipolar interactions originating from the molecular motion in the crystalline domain. The increase in molecular mobility in the crystalline domain was clearly shown by VT T1rho(F) and 1H --> 19F CP measurements in the phase transition temperature range.     

Hydrogen bonding structure and stability of alpha-chitin studied by 13C solid-state NMR

The structure and stability of hydrogen bonds in alpha-chitin were investigated by (13)C solid-state NMR measurements at different temperatures. Splitting of the carbonyl carbon signal for alpha-chitin was interpreted as two types of hydrogen bonding; the peaks at 173.5 and 175.8 ppm were assigned, respectively, to a carbonyl carbon hydrogen bonded exclusively to the NH group and a carbonyl carbon hydrogen-bonded to both NH and C(6)-OH groups. Approximately 60% of carbonyl groups exclusively contributed to the intermolecular hydrogen bonding and ca. 40% of them to the combination of intermolecular and intramolecular hydrogen bonding. Internal rotation around the C(5)-C(6) bond was detected at 55 degrees C.     

Fingerprints of Phalaenopsis Tissues in Growth and Spike Induction Periods—A Solid‐state 13C NMR Approach

Solid‐state Nuclear Magnetic Resonance (ss‐NMR) ¹³C single‐pulse excitation spectroscopy in combination with the magic‐angle spinning (MAS) technique was applied to a series of Phalaenopsis tissues, including the leaf, sheath, stem, and root, at different growth and spiking periods. Compared with{¹H}/¹³C cross‐polarization MAS spectra, the ¹³C single‐pulse excitation MAS spectra displayed very distinct spectral patterns, recognizable as fingerprints of the tissues studied. ¹Here, we demonstrate that solid‐state ¹³C single‐pulse excitation NMR spectroscopy provides a direct and robust analytical tool for studying the various tissues of Phalaenopsis in different growth and spiking induction periods.     

Effects of polymorphic differences for sulfanilamide, as seen through 13C and 15N solid-state NMR, together with shielding calculations

We recorded both carbon-13 and nitrogen-15 NMR spectra of the three solid forms of sulfanilamide most commonly known. This study led to an interpretation of the solid-state effects seen in cross-polarization magic angle spinning spectra. Relaxation times for the different forms were measured. These show different behaviour for the three forms, arising from mobility variations. To obtain information on local environments, static spectra and spinning sideband manifolds were recorded and analysed for the 15N resonances, using isotopically enriched samples. Shielding asymmetries and anisotropies for the two nitrogen nuclei were obtained, showing very different behaviour for the two sites. Shielding calculations were carried out for both 13C and 15N nuclei, and the results are discussed in relation to the experimental values.     

Solid-state NMR detection of 14N-13C dipolar couplings between amino acid side groups provides constraints on amyloid fibril architecture

Solid‐state nuclear magnetic resonance (SSNMR) is a powerful technique for the structural analysis of amyloid fibrils. With suitable isotope labelling patterns, SSNMR can provide constraints on the secondary structure, alignment and registration of β‐strands within amyloid fibrils and identify the tertiary and quaternary contacts defining the packing of the β‐sheet layers. Detection of ¹⁴N? ¹³C dipolar couplings may provide potentially useful additional structural constraints on β‐sheet packing within amyloid fibrils but has not until now been exploited for this purpose. Here a frequency‐selective, transfer of population in double resonance SSNMR experiment is used to detect a weak ¹⁴N? ¹³C dipolar coupling in amyloid‐like fibrils of the peptide H₂N‐SNNFGAILSS‐COOH, which was uniformly ¹³C and ¹⁵N labelled across the four C‐terminal amino acids. The ¹⁴N? ¹³C interatomic distance between leucine and asparagine side groups is constrained between 2.4 and 3.8 Å, which allows current structural models of the β‐spine arrangement within the fibrils to be refined. This procedure could be useful for the general structural analysis of other proteins in condensed phases and environments, such as biological membranes. Copyright © 2011 John Wiley & Sons, Ltd.