Using chemical shift anisotropy to resolve isotropic signals in solid-state NMR

A key problem in solid-state NMR is resolving overlapping isotropic signals. We present here a two-dimensional method which can enable sites with the same isotropic chemical shift to be distinguished according to their chemical shift anisotropy and asymmetry. The method involves correlating sideband spectra at different effective spinning rates using CSA-amplification pulse sequences. The resulting two-dimensional correlation pattern allows very accurate determination of the chemical shift principal values in addition to the recovery of parameters for two overlapping patterns which allows the resolution of overlapping signals.     

Chemical shift anisotropy amplification

A new NMR experiment which allows a measurement of the chemical shift anisotropy (CSA) tensor under magic angle spinning (MAS) is described. This correlates a fast MAS spectrum in the omega2 dimension with a sideband pattern in omega1 in which the intensities mimic those for a sample spinning at a fraction of the rate omega r/N, and these sidebands result from an amplification by a factor N of the modulation caused by the CSA. Standard methods can be used to extract the principal tensor components from the omega1 sideband patterns, and the nature of the experiment is such that the use of a large number of t1 increments can be avoided without compromising the resolution of different chemical sites. The new experiment is useful for accurately measuring narrow shift anisotropies.     

Ramped-speed cross polarization MAS NMR

The inverse cubic dependency of the acceleration of a rotor on its diameter allows for mechanical dynamics comparable to spin dynamics in coupled spin systems. Rotor acceleration up to 300 kHz/s was measured. This feature can be used to simplify existing experiments and explore entirely new ones in the study of spin topologies and material properties.     

Chemical shift anisotropy selective inversion

Magic angle spinning (MAS) is used in solid-state NMR to remove the broadening effects of the chemical shift anisotropy (CSA). In this work we investigate a technique that can reintroduce the CSA in order to selectively invert transverse magnetization. The technique involves an amplitude sweep of the radio frequency field through a multiple of the spinning frequency. The selectivity of this inversion mechanism is determined by the size of the CSA. We develop a theoretical framework to describe this process and demonstrate the CSA selective inversion with numerical simulations and experimental data. We combine this approach with cross-polarization (CP) for potential applications in multi-dimensional MAS NMR.     

Homonuclear decoupled 13C chemical shift anisotropy in 13C doubly labeled peptides by selective-pulse solid-state NMR

We describe a new experiment for measuring homonuclear-decoupled anisotropic chemical shift patterns in doubly 13C-labeled compounds under magic-angle spinning. The experiment combines a pair of selective and non-selective 180 degrees pulses to suppress the 13C-13C scalar and dipolar interactions. This is combined with the recently developed SUPER technique to recouple the chemical shift anisotropy. Demonstrations on 13Calpha and 13CO-labeled amino acids and peptides show that accurate chemical shift powder patterns can be obtained. This permits the use of chemical shift anisotropy for conformational studies of suitably extensively 13C-labeled peptides and proteins.     

Small 51V chemical shift anisotropy for LaVO4 from MQMAS and MAS NMR spectroscopy

51V MQMAS NMR of the triple-quantum transitions is shown to be particularly useful in the determination of the sign and magnitude of the chemical shift anisotropy (CSA) parameter delta(sigma)(= delta(iso)-delta(zz)) along with the asymmetry parameter (eta(sigma)) for a vanadium environment with a small CSA and a rather strong quadrupole coupling. This is demonstrated for the orthovanadate LaVO(4) for which 51V magic-angle spinning (MAS) NMR of the central and satellite transitions at 14.1T gives precise values for the quadrupole coupling parameters, however, an ambiguous sign for delta(sigma). The CSA parameters are reliably obtained from analysis of the spinning sidebands observed in a 51V triple-quantum MAS experiment. Combining these data with least-squares analysis of the manifold of spinning sidebands in the single-pulse MAS NMR spectrum results in a precise determination of the magnitudes and relative orientation of the 51V quadrupole coupling and CSA tensors for LaVO(4).     

Variable angle NMR spectroscopy and its application to the measurement of residual chemical shift anisotropy

The successful measurement of anisotropic NMR parameters like residual dipolar couplings (RDCs), residual quadrupolar couplings (RQCs), or residual chemical shift anisotropy (RCSA) involves the partial alignment of solute molecules in an alignment medium. To avoid any influence of the change of environment from the isotropic to the anisotropic sample, the measurement of both datasets with a single sample is highly desirable. Here, we introduce the scaling of alignment for mechanically stretched polymer gels by varying the angle of the director of alignment relative to the static magnetic field, which we call variable angle NMR spectroscopy (VA-NMR). The technique is closely related to variable angle sample spinning NMR spectroscopy (VASS-NMR) of liquid crystalline samples, but due to the mechanical fixation of the director of alignment no sample spinning is necessary. Also, in contrast to VASS-NMR, VA-NMR works for the full range of sample inclinations between 0° and 90°. Isotropic spectra are obtained at the magic angle. As a demonstration of the approach we measure 13C-RCSA values for strychnine in a stretched PDMS/CDCl? gel and show their usefulness for assignment purposes. In this context special care has been taken with respect to the exact calibration of chemical shift data, for which three approaches have been derived and tested.     

Three-dimensional 13C shift/1H-15N coupling/15N shift solid-state NMR correlation spectroscopy.

Triple-resonance experiments capable of correlating directly bonded and proximate carbon and nitrogen backbone sites of uniformly 13C- and 15N-labeled peptides in stationary oriented samples are described. The pulse sequences integrate cross-polarization from 1H to 13C and from 13C to 15N with flip-flop (phase and frequency switched) Lee-Goldburg irradiation for both 13C homonuclear decoupling and 1H-15N spin exchange at the magic angle. Because heteronuclear decoupling is applied throughout, the three-dimensional pulse sequence yields 13C shift/1H-15N coupling/15N shift correlation spectra with single-line resonances in all three frequency dimensions. Not only do the three-dimensional spectra correlate 13C and 15N resonances, they are well resolved due to the three independent frequency dimensions, and they can provide up to four orientationally dependent frequencies as input for structure determination. These experiments have the potential to make sequential backbone resonance assignments in uniformly 13C- and 15N-labeled proteins.     

A thermometer for nonspinning solid-state NMR spectroscopy

We discuss a method to determine temperature in a static NMR experiment from the temperature variation of the lead nitrate peak shift.     

Spectral deconvolution of NMR cross polarization data sets

The COmponent-REsolved (CORE) strategy has been employed, for the first time to solid state NMR spectroscopy. CORE was used to extract two time-dependent spectral components in 24 ²⁹Si{¹H} NMR spectra, recorded on a meso-structured silica material under conditions of cross polarization evolution. No prior assumptions were made about the component bandshapes, which were both found to be skewed to higher chemical shifts. For the silica fragments close to protons this skewness could be rationalized by a distribution of the degree of condensation in the silica network; however, for the other component the non-Gaussian shape was unexpected. We expect that the same strategy could be applied to a range of experiments in solid-state NMR spectroscopy, where spectral distributions or kinetic parameters need to be accurately extracted.     

Heteronuclear polarization transfer by symmetry-based recoupling sequences in solid-state NMR

We demonstrate a new set of methods for transferring spin polarization between different nuclear isotopes in magic-angle-spinning solid-state NMR. The technique employs symmetry-based recoupling sequences on one irradiation channel and a simple sequence of between one and three strong radiofrequency pulses on the second channel. A phase shift of the recoupling sequences is applied at the same time as a pi/2 pulse on the second channel. The trajectory of the transferred polarization may be used to estimate heteronuclear distances. The method is particularly attractive for nuclei with low gyromagnetic ratios or for those experiencing strong anisotropic spin interactions, where conventional Hartmann-Hahn cross-polarization is difficult to apply. We demonstrate the method on 1H-13C, 1H-15N and 19F-109Ag systems.     

Sensitivity and resolution enhancement in solid-state NMR spectroscopy of bicelles

Magnetically aligned bicelles are becoming attractive model membranes to investigate the structure, dynamics, geometry, and interaction of membrane-associated peptides and proteins using solution- and solid-state NMR experiments. Recent studies have shown that bicelles are more suitable than mechanically aligned bilayers for multidimensional solid-state NMR experiments. In this work, we describe experimental aspects of the natural abundance (13)C and (14)N NMR spectroscopy of DMPC/DHPC bicelles. In particular, approaches to enhance the sensitivity and resolution and to quantify radio-frequency heating effects are presented. Sensitivity of (13)C detection using single pulse excitation, conventional cross-polarization (CP), ramp-CP, and NOE techniques are compared. Our results suggest that the proton decoupling efficiency of the FLOPSY pulse sequence is better than that of continuous wave decoupling, TPPM, SPINAL, and WALTZ sequences. A simple method of monitoring the water proton chemical shift is demonstrated for the measurement of sample temperature and calibration of the radio-frequency-induced heating in the sample. The possibility of using (14)N experiments on bicelles is also discussed.     

Multidimensional solid-state NMR spectroscopy of plant cell walls

Plant biomass has become an important source of bio-renewable energy in modern society. The molecular structure of plant cell walls is difficult to characterize by most atomic-resolution techniques due to the insoluble and disordered nature of the cell wall. Solid-state NMR (SSNMR) spectroscopy is uniquely suited for studying native hydrated plant cell walls at the molecular level with chemical resolution. Significant progress has been made in the last five years to elucidate the molecular structures and interactions of cellulose and matrix polysaccharides in plant cell walls. These studies have focused on primary cell walls of growing plants in both the dicotyledonous and grass families, as represented by the model plants Arabidopsis thaliana, Brachypodium distachyon, and Zea mays. To date, these SSNMR results have shown that 1) cellulose, hemicellulose, and pectins form a single network in the primary cell wall; 2) in dicot cell walls, the protein expansin targets the hemicellulose-enriched region of the cellulose microfibril for its wall-loosening function; and 3) primary wall cellulose has polymorphic structures that are distinct from the microbial cellulose structures. This article summarizes these key findings, and points out future directions of investigation to advance our fundamental understanding of plant cell wall structure and function.     

Correlating fast and slow chemical shift spinning sideband patterns in solid-state NMR

An experiment is presented that enables the measurement of small chemical shift anisotropy tensors under fast magic-angle spinning (MAS). The two-dimensional spectra obtained give a fast MAS sideband pattern in the directly observed dimension with the spinning sideband intensities equivalent to the chemical shift anisotropy scaled by a factor of N, or equivalently the sample spinning frequency scaled by 1/N, in the indirectly observed dimension. The scaling factor may be arbitrarily varied by changing the number and timings of the rotor synchronized pi-pulses used. Desirable features of the experiment include a fixed length pulse sequence and efficient sampling of the indirectly observed dimension. In addition, neither quadrature detection in the indirect dimension nor storage periods are required, consequently no signal intensity is discarded by the pulse sequence. The experiment is demonstrated using (31)P NMR of sodium phosphate and (13)C NMR of fumaric acid monoethyl ester for which a scaling factor of N=10.2 was employed.     

Symmetry-based recoupling of proton chemical shift anisotropies in ultrahigh-field solid-state NMR

A two-dimensional NMR experiment for estimating proton chemical shift anisotropies (CSAs) in solid powders under magic-angle spinning conditions is demonstrated in which 1H CSAs are reintroduced with a symmetry-based recoupling sequence while the individual proton sites are resolved according to their isotropic chemical shifts by magic-angle spinning (MAS) or combined rotation and multiple pulse (CRAMPS) homonuclear decoupling. The experiments where carried out on an ultrahigh-field solid-state NMR instrument (900 MHz 1H frequency) which leads to increased resolution and reliability of the measured 1H CSAs. The experiment is expected to be important for investigating hydrogen bonding in solids.     

Low temperature probe for dynamic nuclear polarization and multiple-pulse solid-state NMR

Here, we describe the design and performance characteristics of a low temperature probe for dynamic nuclear polarization (DNP) experiments, which is compatible with demanding multiple-pulse experiments. The competing goals of a high-Q microwave cavity to achieve large DNP enhancements and a high efficiency NMR circuit for multiple-pulse control lead to inevitable engineering tradeoffs. We have designed two probes-one with a single-resonance RF circuit and a horn-mirror cavity configuration for the microwaves and a second with a double-resonance RF circuit and a double-horn cavity configuration. The advantage of the design is that the sample is in vacuum, the RF circuits are locally tuned, and the microwave resonator has a large internal volume that is compatible with the use of RF and gradient coils.