Distance measurements between carbon and bromine using a split-pulse PM-RESPDOR solid-state NMR experiment.

Solid-state nuclear magnetic resonance has long been used to probe atomic distances between nearby nuclear spins by virtue of the dipolar interaction. New technological advances have recently enabled simultaneous tuning of the radio-frequency resonance circuits to nuclei with close Larmor frequencies, bringing great promise, among other experiments, also to distance measurements between such nuclei, in particular for nuclei with a spin larger than one-half. However, this new possibility has also required modifications of those experiments since the two nuclei cannot be irradiated simultaneously. When measuring distances between a spin S = 1/2 and a quadrupolar spin (S > ½), this drawback can be overcome by splitting the continuous-wave recoupling pulse applied to the quadrupolar nucleus. We show here that a similar adjustment to a highly-efficient phase-modulated (PM) recoupling pulse enables distance measurements between nuclei with close Larmor frequencies, where the coupled spin experiences a very large coupling. Such an experiment, split phase-modulated RESPDOR, is demonstrated on a 13C-81Br system, where the difference in Larmor frequencies is only 7%, or 11.2 MHz on a 14.1 T magnet. The inter-nuclear distances are extracted using an unscaled analytical formula. Since bromine usually experiences particularly high quadrupolar couplings, as in the current case, we suggest that the split-PM-RESPDOR experiment can be highly beneficial for research on bromo-compounds, including many pharmaceuticals, where carbon-bromine bonds are prevalent, and organo-catalysts utilizing the high reactivity of bromides. We show that for butyl triphenylphosphonium bromide, the solid-state NMR distances are in agreement with a low-hydration compound rather than a water-caged semi-clathrate form. The split-PM-RESPDOR experiment is suitable for distance measurements between any quadrupolar ↔ spin-1/2 pair, in particular when the quadrupolar spin experiences a significantly large coupling.

Pushing the limit of NMR-based distance measurements - retrieving dipolar couplings to spins with extensively large quadrupolar frequencies.

Dipolar recoupling under magic-angle spinning allows to measure accurate inter-nuclear distances provided that the two interacting spins can be efficiently and uniformly excited. Alexander (Lex) Vega has shown that adiabatic transfers of populations in quadrupolar spins during the application of constant-wave (cw) radio-frequency pulses lead to efficient and quantifiable dipolar recoupling curves. Accurate distance determination within and beyond the adiabatic regime using cw pulses is limited by the size of the quadrupolar coupling constant. Here we show that using the approach of long-pulse phase modulation, dipolar recoupling and accurate distances can be obtained for nuclei having extensively large quadrupolar frequencies of 5-10 MHz. We demonstrate such results by obtaining a 31P-79/81Br distance in a compound for which bromine-79 (spin-3/2) has a quadrupolar coupling constant of 11.3 MHz, and a 13C-209Bi distance where the bismuth (spin-9/2) has a quadrupolar coupling constant of 256 MHz, equaling a quadrupolar frequency of 10.7 MHz. For Bromine, we demonstrate that an analytical curve based on the assumption of complete spin saturation fits the data. In the case of bismuth acetate, a C-Bi3 spin system must be used in order to match the correct saturation recoupling curve, and results are in agreement with the crystallographic structure.

Analysis of large-anisotropy-spin recoupling pulses for distance measurement under magic-angle spinning NMR shows the superiority and robustness of a phase modulated saturation pulse.

The distance between a spin one-half and an attached spin possessing a large anisotropy can be obtained using different dipolar recoupling sequences that are based on the rotational-echo double resonance technique under magic-angle spinning solid-state NMR. The general difference between these sequences with respect to the coupled spin is the set of pulses applied in order to drive this spin out of equilibrium, thereby recoupling the dipolar interaction. Since complete inversion is practically not possible due to the coupled-spin anisotropy, using one or another pulse depends on the experimental and spin conditions: the spinning speed, the strength of the radio frequency field, the size of the anisotropic interaction (quadrupolar or chemical shiftanisotropy couplings), the offset, and the accuracy of setting the magic angle. Here we present a detailed description of the behavior of the anisotropic spin magnetization, including the macroscopic level transition probabilities, the degree of inversion, and the microscopic and macroscopic magnetizations during the applications of these pulses under different experimental conditions. As simulations show, a complete randomization of spin populations under a wide range of experimental conditions occurs under a specific phase modulation of the recoupling pulse while for all other cases dependence on experimental conditions is large and the achievable bandwidth is limited. A result of this detailed analysis is that the extension of the phase modulated pulse extends even further its robustness. The saturation capability is demonstrated experimentally for the quadrupolar spin of boron-11 in 4-methoxyphenylboronic acid.

Site-resolved multiple-quantum filtered correlations and distance measurements by magic-angle spinning NMR: Theory and applications to spins with weak to vanishing quadrupolar couplings.

We discuss and analyze four magic-angle spinning solid-state NMR methods that can be used to measure internuclear distances and to obtain correlation spectra between a spin I = 1/2 and a half-integer spin S > 1/2 having a small quadrupolar coupling constant. Three of the methods are based on the heteronuclear multiple-quantum and single-quantum correlation experiments, that is, high rank tensors that involve the half spin and the quadrupolar spin are generated. Here, both zero and single-quantum coherence of the half spins are allowed and various coherence orders of the quadrupolar spin are generated, and filtered, via active recoupling of the dipolar interaction. As a result of generating coherence orders larger than one, the spectral resolution for the quadrupolar nucleus increases linearly with the coherence order. Since the formation of high rank tensors is independent of the existence of a finite quadrupolar interaction, these experiments are also suitable to materials in which there is high symmetry around the quadrupolar spin. A fourth experiment is based on the initial quadrupolar-driven excitation of symmetric high order coherences (up to p = 2S, where S is the spin number) and subsequently generating by the heteronuclear dipolar interaction higher rank (l + 1 or higher) tensors that involve also the half spins. Due to the nature of this technique, it also provides information on the relative orientations of the quadrupolar and dipolar interaction tensors. For the ideal case in which the pulses are sufficiently strong with respect to other interactions, we derive analytical expressions for all experiments as well as for the transferred echo double resonance experiment involving a quadrupolar spin. We show by comparison of the fitting of simulations and the analytical expressions to experimental data that the analytical expressions are sufficiently accurate to provide experimental (7)Li-(13)C distances in a complex of lithium, glycine, and water. Discussion of the regime for which such an approach is valid is given.

An optimal double-magic flip angle for performing the distance measurement REDOR experiment on a spin S=1.

Distance measurements between a half-spin and a quadrupolar S=1 spin having a small quadrupolar coupling constant can be performed using the rotational echo double resonance (REDOR) experiment. We derived an analytical expression for the probability of transitions between energy levels resulting from the application of an arbitrary pulse flip angle to the quadrupolar spin and consequently minimized the probability that populations of individual levels do not undergo a spin transition during the pulse. As a result we discovered that if the flip angle of the quadrupolar spin pulse is 109.47°, the maximal recoupling values are the largest possible and the signal reaches a maximum value of 8/9, larger than in the use of either a 90° pulse or a 180° pulse. In addition, the slope of the initial decay is higher than that of the 90° pulse. The recoupling signal can be modeled by an exact analytical formula in the ideal case and simulations show that the advantage of the 109.47° pulse is preserved when the quadrupolar coupling constant CQ has a finite value typical of (2)H and (6)Li spins (up to CQ~200kHz). Experimental results on two spin pairs, (2)H-(13)C and (6)Li-(13)C, demonstrate the validity and accuracy of this method.

Shimon Vega in the eyes of his students and postdocs.

Professor Shimon Vega (1943-2021) of the Weizmann Institute of Science passed away on the 16-th of November. Shimon Vega established theoretical frameworks to develop and explain solid-state nuclear magnetic resonance (NMR) and dynamic nuclear polarization (DNP) techniques and methodologies. His departure left a profound mark on his many students, postdocs, and colleagues. Shortly after his passing, we all assembled spontaneously for an international online meeting to share our reflections and memories of our experiences in Shimon's lab and how they affected us deeply during that period of timeand throughout our scientific careers. These thoughts and feelings were put here into writing.

How does the mood stabilizer lithium bind ATP, the energy currency of the cell: Insights from solid-state NMR.

BACKGROUND: Lithium, in the form of a salt, is a mood stabilizer and a leading drug for the treatment of bipolar disorder. It has a very narrow therapeutic range and a variety of side effects. Lithium can replace magnesium and other cations in enzymes and small molecules, among them ATP, thereby affecting and inhibiting many biochemical pathways. The form of binding of lithium ions to ATP is not known. METHODS: Here we extract the binding environment of lithium in solid ATP using a multi-nuclear multi-dimensional solid-state NMR approach. RESULTS: We determine that the coordination sphere of lithium includes, at a distance of 3.0(±0.4) Å, three phosphates; the two phosphates closest to the ribose ring from one ATP molecule, and the middle phosphate from another ATP molecule. A water molecule most probably completes the fourth coordination. Despite the use of excess lithium in the preparations, sodium ions still remain bound to the sample, at distances of 4.3-5.5 Šfrom Li, and coordinate the first phosphate and two terminal phosphates. CONCLUSIONS: Solid-state NMR enables to unravel the exact coordination of lithium in ATP showing binding to three phosphates from two molecules, none of which are the terminal gamma phosphate. GENERAL SIGNIFICANCE: The methods we use are applicable to study lithium bound to a variety of ATP-bound enzymes, or to other cellular targets of lithium, consequently suggesting a molecular basis for its mode of action.

Characterization of lithium coordination sites with magic-angle spinning NMR.

Lithium, in the form of lithium carbonate, is one of the most common drugs for bipolar disorder. Lithium is also considered to have an effect on many other cellular processes hence it possesses additional therapeutic as well as side effects. In order to quantitatively characterize the binding mode of lithium, it is required to identify the interacting species and measure their distances from the metal center. Here we use magic-angle spinning (MAS) solid-state NMR to study the binding site of lithium in complex with glycine and water (LiGlyW). Such a compound is a good enzyme mimetic since lithium is four-coordinated to one water molecule and three carboxylic groups. Distance measurements to carbons are performed using a 2D transferred echo double resonance (TEDOR) MAS solid-state NMR experiment, and water binding is probed by heteronuclear high-resolution proton-lithium and proton-carbon correlation (wPMLG-HETCOR) experiments. Both HETCOR experiments separate the main complex from impurities and non-specifically bound lithium species, demonstrating the sensitivity of the method to probe the species in the binding site. Optimizations of the TEDOR pulse scheme in the case of a quadrupolar nucleus with a small quadrupole coupling constant show that it is most efficient when pulses are positioned on the spin-1/2 (carbon-13) nucleus. Since the intensity of the TEDOR signal is not normalized, careful data analysis that considers both intensity and dipolar oscillations has to be performed. Nevertheless we show that accurate distances can be extracted for both carbons of the bound glycine and that these distances are consistent with the X-ray data and with lithium in a tetrahedral environment. The lithium environment in the complex is very similar to the binding site in inositol monophosphatase, an enzyme associated with bipolar disorder and the putative target for lithium therapy. A 2D TEDOR experiment applied to the bacterial SuhB gene product of this enzyme was designed to probe direct correlations between lithium, the enzyme inhibitor, and the closest carboxyl carbons of the binding site. At this point, the chemical shift of the bound carboxyl groups in this 29 kDa enzyme could be determined.

High resolution heteronuclear correlation NMR spectroscopy between quadrupolar nuclei and protons in the solid state.

A high resolution two-dimensional solid state NMR experiment is presented that correlates half-integer quadrupolar spins with protons. In this experiment the quadrupolar nuclei evolve during t1 under a split-t1, FAM-enhanced MQMAS pulse scheme. After each t1 period ending at the MQMAS echo position, single quantum magnetization is transferred, via a cross polarization process in the mixing time, from the quadrupolar nuclei to the protons. High-resolution proton signals are then detected in the t2 time domain during wPMLG5* homonuclear decoupling. The experiment has been demonstrated on a powder sample of sodium citrate and 23Na-1H 2D correlation spectra have been obtained. From the HETCOR spectra and the regular MQMAS spectrum, the three crystallographically inequivalent Na+ sites in the asymmetric unit were assigned. This MQMAS-wPMLG HETCOR pulse sequence can be used for spectral editing of half-integer quadrupolar nuclei coupled to protons.

Interatomic distance measurement in solid-state NMR between a spin- 1/2 and a spin- 5/2 using a universal REAPDOR curve.

A universal curve for the solid-state NMR REAPDOR experiment on an isolated spin-1/2-spin-5/2 pair is proposed that provides a simple means to measure their interatomic distance. REAPDOR data were obtained at three separate REAPDOR experiments using different values of the rotor spinning frequency. All points were fitted simultaneously to the universal formula without a need for full density matrix calculations. The 13C-17O distance of 2.45 A was measured between the C6 carbon and the 17O label in a tyrosine sample. The error of 8% in the dipolar coupling (Dfit = 278 Hz) is well within the 15% theoretical tolerance of this curve.

The influence of the radiofrequency excitation and conversion pulses on the lineshapes and intensities of the triple-quantum MAS NMR spectra of I = 3/2 nuclei.

A rigorous examination of the various multiple-quantum magic angle spinning sequences is carried out with reference to sensitivity enhancement in the isotropic dimension and the lineshapes of the corresponding MAS peaks in the anisotropic dimension. An echo efficiency parameter is defined here, which is shown to be an indicator of the performance aspects of the various sequences. This can be used in the design of further new experiments in this field. A consequence of such a systematic analysis has been the combination of a spin-lock pulse for excitation of multiple-quantum coherences and an amplitude-modulated pulse for their conversion to observable single-quantum coherences. This approach has resulted in an improved performance over other sequences with respect to both the anisotropic lineshapes and the isotropic intensities.

Deuterium REDOR: principles and applications for distance measurements

The application of short composite pulse schemes ( and ) to the rotational echo double-resonance (REDOR) spectroscopy of X-2H (X: spin 12, observed) systems with large deuterium quadrupolar interactions has been studied experimentally and theoretically and compared with simple 180 degrees pulse schemes. The basic properties of the composite pulses on the deuterium nuclei have been elucidated, using average Hamiltonian theory, and exact simulations of the experiments have been achieved by stepwise integration of the equation of motion of the density matrix. REDOR experiments were performed on 15N-2H in doubly labeled acetanilide and on 13C-2H in singly 2H-labeled acetanilide. The most efficient REDOR dephasing was observed when composite pulses were used. It is found that the dephasing due to simple 180 degrees deuterium pulses is about a factor of 2 less efficient than the dephasing due to the composite pulse sequences and thus the range of couplings observable by X-2H REDOR is enlarged toward weaker couplings, i.e., larger distances. From these experiments the 2H-15N dipolar coupling between the amino deuteron and the amino nitrogen and the 2H-13C dipolar couplings between the amino deuteron and the alpha and beta carbons have been elucidated and the corresponding distances have been determined. The distance data from REDOR are in good agreement with data from X-ray and neutron diffraction, showing the power of the method. Copyright 1999 Academic Press.