Two-dimensional Solid-state NMR Experiments Research Articles

Glasses in the NaPO3-WO3-NaF ternary system: preparation, physical properties and structural studies

Oxyfluoride glasses with composition x WO3 - 30 NaPO3 - (70-x) NaF, with 30 ≤ x ≤ 70 were prepared by melt-quenching and characterized with respect to their bulk physical properties and ionic conductivities. As the NaF content increases, the glass transition temperature, Tg, decreases systematically. Impedance measurements reveal no clear effect of NaF content upon room temperature electrical conductivities and activation energies. There is, however, a significant decrease of the Arrhenius pre-exponential factor with increasing WO3 content, suggesting that the electrical conductivities measured in this glass series are increasingly influenced by an electronic contribution. This contribution may be related to a W5+ → W6+ electron hopping process detected in the optical absorption spectra. Detailed spectroscopic investigations by Raman scattering and multinuclear one- and two-dimensional solid-state NMR experiments indicate that NaF acts like a network modifier resulting in the formation of W-F bonds via breakage of W-O-W and W-O-P linkages, with the former process being the preferred one. The concomitant decrease in network connectivity caused by these processes accounts for the experimentally observed decrease in glass transition temperatures with increasing NaF contents.

Three pulse recoupling and phase jump matching

The paper describes a family of novel recoupling pulse sequences, called three pulse recoupling. These pulse sequences can be employed for both homonuclear and heteronuclear recoupling experiments and are robust to dispersion in chemical shifts and rf-inhomogeneity. These recoupling pulse sequences can be used in design of two-dimensional solid state NMR experiments that use powdered dephased antiphase coherence (γ preparation) to encode chemical shifts in the indirect dimension. Both components of this chemical shift encoded gamma-prepared states can be refocused into inphase coherence by a recoupling element. This helps to achieve sensitivity enhancement in 2D NMR experiments by quadrature detection.

The use of a selective saturation pulse to suppress t1 noise in two-dimensional 1H fast magic angle spinning solid-state NMR spectroscopy

A selective saturation pulse at fast magic angle spinning (MAS) frequencies (60+kHz) suppresses t1 noise in the indirect dimension of two-dimensional 1H MAS NMR spectra. The method is applied to a synthetic nucleoside with an intense methyl 1H signal due to triisopropylsilyl (TIPS) protecting groups. Enhanced performance in terms of suppressing the methyl signal while minimising the loss of signal intensity of nearby resonances of interest relies on reducing spin diffusion – this is quantified by comparing two-dimensional 1H NOESY-like spin diffusion spectra recorded at 30–70kHz MAS. For a saturation pulse centred at the methyl resonance, the effect of changing the nutation frequency at different MAS frequencies as well as the effect of changing the pulse duration is investigated. By applying a pulse of duration 30ms and nutation frequency 725Hz at 70kHz MAS, a good compromise of significant suppression of the methyl resonance combined with the signal intensity of resonances greater than 5ppm away from the methyl resonance being largely unaffected is achieved. The effectiveness of using a selective saturation pulse is demonstrated for both homonuclear 1H–1H double quantum (DQ)/single quantum (SQ) MAS and 14N–1H heteronuclear multiple quantum coherence (HMQC) two-dimensional solid-state NMR experiments.

Novel expression and characterization of a light driven proton pump archaerhodopsin 4 in a Halobacterium salinarum strain

Archaerhodopsin 4 (AR4), a new member of the microbial rhodopsin family, is isolated from Halobacterium species xz515 in a Tibetan salt lake. AR4 functions as a proton pump similar to bacteriorhodopsin (BR) but with an opposite temporal order of proton uptake and release at neutral pH. However, further studies to elucidate the mechanism of the proton pump and photocycle of AR4 have been inhibited due to the difficulty of establishing a suitable system in which to express recombinant AR4 mutants. In this paper, we report a reliable method for expressing recombinant AR4 in Halobacterium salinarum L33 with a high yield of up to 20mg/l. Experimental results show that the recombinant AR4 retains the light-driven proton pump characteristics and photo-cycling kinetics, similar to that in the native membrane. The functional role of bacterioruberin in AR4 and the trimeric packing of AR4 in its native and recombinant forms are investigated through light-induced kinetic measurements, two-dimensional solid-state NMR experiments, dynamic light scattering (DLS) and Fourier transform infrared spectroscopy (FTIR). Such approaches provide new insights into structure–function relationships of AR4, and form a basis for other archaeal rhodopsins.

Indirect “No-Bond” 31P···31P Spin–Spin Couplings in P,P-[3]Ferrocenophanes: Insights from Solid-State NMR Spectroscopy and DFT Calculations

No-bond (31)P-(31)P indirect dipolar couplings, which arise from the transmission of nuclear spin polarization through interaction of proximal nonbonded electron pairs have been investigated in the solid state for a series of closely related substituted P,P-[3]ferrocenophanes and model systems. Through variation and combination of ligands (phenyl, cyclohexyl, isopropyl) at the two phosphorus sites, the P···P distances in these compounds can be varied from 3.49 to 4.06 Å. Thus, the distance dependence of the indirect no-bond coupling constant J(nb) can be studied in a series of closely related compounds. One- and two-dimensional solid-state NMR experiments serve to establish the character of these couplings and to measure the isotropic coupling constants J(iso), which were found to range between 12 and 250 Hz. To develop an understanding of the magnitude of J(nb) in terms of molecular structure, their dependences on intramolecular internuclear distances and relative orbital orientations is discussed by DFT-calculations on suitable models. In agreement with the literature the dependence of J(nb) on the P···P distance is found to be exponential; however, the steepness of this curve is highly dependent on the internuclear equilibrium distance. For a quantitative description, the off-diagonal elements of the expectation value of the Kohn-Sham-Fock operator in the LMO basis for the LMOs of the two phosphorus lone-pairs is proposed. This parameter correlates linearly with the calculated J(nb) values and possesses the same distance-dependence. In addition, the simulations indicate a distinct dependence of J(nb) on the dihedral angle defined by the two C-P bonds providing ligation to the molecular backbone. For disordered materials or those featuring multiple sites, conformers, and/or polymorphism, a new double-quantum NMR method termed DQ-DRENAR can be used to conveniently measure internuclear (31)P-(31)P distances. If conducted on compounds with known P···P distances, such measurements can also serve to estimate the magnitude of the anisotropy ΔJ of these no-bond indirect spin-spin couplings. The DFT results suggest that in the present series of compounds the magnitude of ΔJ is strongly correlated with that of the isotropic component, as both parameters have analogous distance dependences. While our studies indicate a sizable J-anisotropy for the model compound 1,8-bis(diphenylphosphino)napthalene (ΔJ ~ -70 Hz), the corresponding values for the P,P-[3]ferrocenophanes are significantly smaller, affecting their DQ-DRENAR curves only in a minor way. Based on the above insights, the structural aspects of conformational disorder and polymorphism observed in some of the P,P-[3]ferrocenophanes are discussed.

Mineral Association Changes the Secondary Structure and Dynamics of Murine Amelogenin

Amelogenin is one of the key protein constituents responsible for the exquisite organization of the calcium phosphate crystals in enamel. Amelogenin forms into nanospheres in solution, while its association with hydroxyapatite is also essential to enamel development. Structural information of full-length amelogenin in either of these physiologically important forms has the potential to provide mechanistic information; however, these data are limited because of the difficulty of determining the structure of large protein complexes and proteins bound to surfaces. To obtain structural insights into amelogenin during these early stages of enamel development, we used a lysine-specific 13C-, 15N-labeled sample of murine amelogenin to provide insight into the structure of the hydroxyapatite (HAP)-binding domains of the protein. A combination of one-and two-dimensional solid-state NMR experiments was used to obtain molecular-level insights into the secondary structure and dynamics of full-length amelogenin within a nanosphere-gel and on the surface of HAP. Regions of amelogenin that appear to be primarily random coil in the nanosphere-gel adopt a β-strand structure and are less mobile with HAP binding, indicative of a structural switch upon binding that may be important in the role of amelogenin in enamel development.

Recoupling in solid state NMR using γ prepared states and phase matching

The paper describes two-dimensional solid state NMR experiments that use powdered dephased antiphase coherence ( γ preparation) to encode chemical shifts in the indirect dimension. Both components of this chemical shift encoded gamma-prepared states can be refocused into inphase coherence by a recoupling element. This helps to achieve sensitivity enhancement in 2D NMR experiments by quadrature detection. The powder dependence of the gamma-prepared states allows for manipulating them by suitable insertion of delays in the recoupling periods. This helps to design experiments that suppress diagonal peaks in 2D spectra, leading to improved resolution. We describe some new phase modulated heteronuclear and homonuclear recoupling pulse sequences that simplify the implementation of the described experiments based on γ prepared states. Recoupling in the heteronuclear spin system is achieved by matching the difference in the amplitude of the sine/cosine modulated phase on the two rf-channels to the spinning frequency while maintaining the same power on the two rf-channels.

Structure of Amorphous Aluminum Oxide

Whereas prototypical Al(2)O(3) is not a glass former, amorphous Al(2)O(3) can be formed as thin films through vapor deposition and can serve as a structural model for the Al(2)O(3) glass. The first two-dimensional solid-state NMR experiments for amorphous Al(2)O(3) thin film reveal that four- and five-coordinated species are predominant (95%), while six-coordinated species are minor. Such a species distribution is remarkably similar to what has been predicted theoretically for Al(2)O(3) melts. Upon annealing to 800 degrees C the five-coordinated species becomes negligible, indicating the onset of crystallization of Al(2)O(3).

Magic angle spinning (MAS) NMR: a new tool to study the spatial and electronic structure of photosynthetic complexes

In the last two decades, Magic Angle Spinning (MAS) NMR has created its own niche in studies involving photosynthetic membrane protein complexes, owing to its ability to provide structural and functional information at atomic resolution of membrane proteins when in the membrane, in the natural environment. The light-harvesting two (LH2) transmembrane complex from Rhodopseudomonas acidophila is used to illustrate the procedure of the technique applicable in photosynthesis research. One- and two-dimensional solid-state NMR experiments involving 13C- and 15N-labeled LH2 complexes allow to make a sequence-specific assignment of NMR signals, which forms the basis for resolving structural details and the assessment of charge transfer, electronic delocalization effects, and functional strain in the ground state.

Formation and Characterization of Crystals Containing a Pleuromutilin Derivative, Succinic Acid and Water

An unusual pharmaceutical solid containing a pleuromutilin derivative, succinic acid and water is studied with a combination of physiochemical methods. The parent pleuromutilin derivative creates a host structure that contains large channels, which are occupied by variable amounts of both succinic acid and water. The inherent disorder of the system does not lend itself to detailed single-crystal X-ray diffraction studies; hence a combination of powder X-ray diffraction (PXRD) and solid-state NMR (SSNMR) techniques were used to characterize this system and understand its behavior. A variety of one and two-dimensional SSNMR experiments were used to understand the proton transfer and the hydrogen bonding network, including 1H−13C heteronuclear correlation experiments and measurements of the principal components of 13C chemical shift tensors. This crystal system can exist as two distinct phases; a hemisuccinate phase containing a doubly ionized succinate molecule with two cations of the pleuromutilin derivati...

Probing membrane protein orientation and structure using fast magic-angle-spinning solid-state NMR

One and two-dimensional solid-state NMR experiments are discussed that permit probing local structure and overall molecular conformation of membrane-embedded polypeptides under Magic Angle Spinning. The functional dependence of a series of anisotropic recoupling schemes is analyzed using theoretical and numerical methods. These studies lead to the construction of a set of polarization dephasing or transfer units that probe local backbone conformation and overall molecular orientation within the same NMR experiment. Experimental results are shown for a randomly oriented peptide and for two model membrane-peptides reconstituted into lipid bilayers and oriented on polymer films according to a method proposed by Bechinger et al.

Self-Organization, Structure, Dynamic Properties, and Surface Morphology of Silica/Epoxy Films As Seen by Solid-State NMR, SAXS, and AFM

The high degree of self-assembling of epoxide-based products made from functionalized organosilica building blocks, functionalized oligo(oxypropylene)-diamine and/or -triamine, and colloidal silica nanoparticles was evidenced by solid-state NMR spectroscopy, small-angle X-ray scattering (SAXS), and atomic force microscopy (AFM). Under optimized conditions of preparation, isolated siloxane cagelike clusters arise in the reaction mixture. No cleavage of oxirane rings occurs before thermal curing, and thus the whole process is well controlled. The presence of silica nanoparticles accelerates the kinetics of polycondensation and affects the condensation rate of siloxane units in final products. Two-dimensional solid-state NMR experiments (2D CRAMPS, 2D 1H−13C and 1H−29Si HETCOR, WISE) revealed differences in structure and segmental dynamics of final films as well as in self-organization and homogeneity degree depending on reaction conditions. Ideally, siloxane cagelike clusters are regularly dispersed within the bulk and oxypropylene chains form phase which separates organic tails of siloxane clusters. The SAXS determined distance between clusters (1.8 nm) well corresponds to the constraints determined by NMR spin-diffusion experiments. Polymer interaction with silica nanoparticles is confirmed by two-dimensional 1H−29Si HETCOR experiments.

Vacancy ordering and host–guest interactions in CdPS3 intercalates: Results from multidimensional solid state NMR

Cadmium hexathiohypodiphosphate (CdPS3) intercalates with potassium and N,N′-dimethylviologen cations have been studied by complementary one- and two-dimensional solid state NMR experiments. The intercalation process results in vacancies in the Cd sublattice, whose detailed distribution is sensitively reflected in both the 31P and the 113Cd spectra of the host material. Advanced 2-D 31P double quantum and spin echo NMR techniques are used to further investigate the connectivity patterns of the 31P atoms represented by the peaks observed. Simple chemical shift considerations strongly indicate that the vacancy distribution is primarily controlled by electrostatic repulsion effects. Thus a model is developed for the local vacancy arrangement that links structural motifs with stoichiometry and the NMR results. In the case of the methylviologen intercalate NMR allows the study of the interaction between the organic guest molecule and the host material. Owing to strong Coulombic interactions, one positively charged end of the molecule is pinned at a minimum distance to the negatively charged Cd vacancy in the layer on the NMR time scale, while the other end is orientationally disordered relative to the layers. This arrangement provides an excellent fit to the experimental intensity distributions in the one- and two-dimensional 31P MAS NMR spectra.

Structure and topology of a peptide segment of the 6th transmembrane domain of the Saccharomyces cerevisae alpha-factor receptor in phospholipid bilayers.

A detailed analysis of the structure of an 18-residue peptide AQSLLVPSIIFILAYSLK [M6(252-269, C252A)] in 1,2-dimyristoyl-sn-glycero-phosphocholine bilayers was carried out using solid state NMR and attenuated total reflection Fourier transform infrared spectroscopy. The peptide corresponds to a portion of the 6th transmembrane domain of the alpha-factor receptor of Saccharomyces cerevisiae. Ten homologs of M6(252-269, C252A) were synthesized in which individual residues were labeled with (15)N. One- and two-dimensional solid state NMR experiments were used to determine the chemical shifts and (1)H-(15)N dipolar coupling constants for the (15)N-labeled peptides in oriented dimyristoylphosphatidylcholine bilayers on stacked glass plates. These parameters were used to calculate the structure and orientation of M6(252-269, C252A) in the bilayers. The results indicate that the carboxyl terminal residues (9-14) are alpha-helical and oriented with an angle of about 8 degrees with respect to the bilayer normal. Independently, an attenuated total reflection Fourier transform infrared spectroscopy analysis on M6(252-269, C252A) in a 1,2-dimyristoyl-sn-glycero-phosphocholine bilayer concluded that the helix tilt angle was about 12.5 degrees. The results on the structure of M6(252-269, C252A) in bilayers are in good agreement with the structure determined in trifluoroethanol/water solutions (B. Arshava et al. Biopolymers, 1998, Vol. 46, pp. 343-357). The present study shows that solid state NMR spectroscopy can provide high resolution information on the structure of transmembrane domains of a G protein-coupled receptor.

Heteronuclear 2D-correlations in a uniformly [13C, 15N] labeled membrane-protein complex at ultra-high magnetic fields.

One- and two-dimensional solid-state NMR experiments on a uniformly labeled intrinsic membrane-protein complex at ultra-high magnetic fields are presented. Two-dimensional backbone and side-chain correlations for a [U-13C, 15N] labeled version of the LH2 light-harvesting complex indicate significant resolution at low temperatures and under Magic Angle Spinning. Tentative assignments of some of the observed correlations are presented and attributed to the alpha-helical segments of the protein, mostly found in the membrane interior.

Conformation and Dynamics of Atactic Poly(acrylonitrile). 1. Trans/Gauche Ratio from Double-Quantum Solid-State 13C NMR of the Methylene Groups

The trans/gauche ratio of backbone bonds in atactic poly(acrylonitrile) (aPAN) was characterized by several two-dimensional solid-state NMR experiments, using 15% 13CH2-carbon labeled aPAN. An estimate of the conformational ratio was obtained from a double-quantum filtered NMR experiment, which correlates the 13C−13C internuclear vector of two adjacent 13CH2 groups with their 13C chemical shift anisotropies. The orientation of the chemical shift tensor was determined in auxiliary experiments. The backbone conformation of two successive bonds was characterized by double-quantum NMR spectroscopy, which correlates the orientation-dependent 13C chemical shifts of two adjacent 13CH2 segments. The experiments were performed with and without 13C−13C dipolar decoupling. From the three two-dimensional NMR spectra that were obtained, the trans/gauche ratio of aPAN was determined as 90%:10% (±10%). This value is compatible with the previously reported experimental density and wide-angle X-ray diffraction patterns of...

Self-Assembled Monolayers of Alkanoic Acids: A Solid-State NMR Study

Long-chain alkanoic acids (CH3(CH3)nCO2H, n > 15) form conformationally ordered monolayers on zirconium oxide powder via a chelating bidentate zirconium carboxylate surface bond. ω-Hydroxyalkanoic acids (HO(CH3)15CO2H) produce hydrophilic monolayers, with no evidence of looping to the metal oxide surface. Variable temperature and two-dimensional solid-state NMR experiments demonstrate that alkanoate monolayers display chain dynamics similar to other self-assembled monolayers (SAMs). The thermal behavior of alkanoate SAMs is also compared to analogous fatty acid salt Langmuir−Blodgett monolayers which undergo pretransitional chain disordering.

A set of two-dimensional solid-state NMR experiments providing topological information for structure refinement of TMP-GaPO

A procedure for structural characterization based on X-ray diffraction (XRD) and on a set of two-dimensional solid-state NMR experiments is proposed. It has been applied to a new gallophosphate oxyfluorinated compound, T(T = amine)—GaPO [Ga4(PO4)3(HPO4)F3·T]. To complement and validate structural information obtained by XRD, a set of three solid-state NMR experiments—RFDR for fluorine-19, double quanta for phosphorus-31 and HETCOR between fluorine and phosphorus—facilitate the analysis of the topology of homoatomic and heteroatomic sub-networks of fluorine and phosphorus included in the inorganic framework of this material. The efficiency of this combination of NMR experiments in assigning all NMR signals to their crystallographically sites is also demonstrated. Copyright © 1999 John Wiley & Sons, Ltd.

Study of phospholipid structure by 1H, 13C, and 31P dipolar couplings from two-dimensional NMR

Various motionally averaged 31P-1H, 13C-1H, 1H-1H, and 31P-13C dipolar couplings were measured for natural-abundance and unoriented phosphocholine in the L alpha phase. The couplings were obtained and assigned by a variety of advanced and partly novel two-dimensional solid-state NMR experiments. Whereas 31P-1H and 31P-13C dipolar couplings provide long-range structural constraints, geminal 1H-1H couplings and the signs of 13C-1H couplings are important new elements in a segmental order-tensor analysis of the lipid headgroup and glycerol backbone. The implications of these measured dipolar couplings for the conformational exchange of the lipid headgroup and the bending of the headgroup from the glycerol backbone are discussed. These dipolar couplings are also analyzed semiquantitatively in terms of the segmental order tensor.

4891596 Magnetic resonance imaging apparatus

The paper describes a family of novel recoupling pulse sequences, called three pulse recoupling. These pulse sequences can be employed for both homonuclear and heteronuclear recoupling experiments and are robust to dispersion in chemical shifts and rf-inhomogeneity. These recoupling pulse sequences can be used in design of two-dimensional solid state NMR experiments that use powdered dephased antiphase coherence (γ preparation) to encode chemical shifts in the indirect dimension. Both components of this chemical shift encoded gamma-prepared states can be refocused into inphase coherence by a recoupling element. This helps to achieve sensitivity enhancement in 2D NMR experiments by quadrature detection.