Fast multidimensional NMR by polarization sharing

The speed of multidimensional NMR spectroscopy can be significantly increased by drastically shortening the customary relaxation delay between scans. The consequent loss of longitudinal magnetization can be retrieved if 'new' polarization is transferred from nearby spins. For correlation spectroscopy involving heteronuclei (X=13C or 15N), protons not directly bound to X can repeatedly transfer polarization to the directly bound protons through Hartmann-Hahn mixing. An order of magnitude increase in speed has been observed for the 600 MHz two-dimensional HMQC spectra of amikacin and strychnine using this technique, and it also reduces the noisy F1 ridges that degrade many heteronuclear correlation spectra recorded with short recovery times.     

Heteronuclear isotropic mixing separated local field NMR spectroscopy

This paper presents a theoretical, numerical, and experimental study of a new class of separated local field (SLF) techniques. These techniques are based on the heteronuclear isotropic mixing leading to spin exchange via the local field (HIMSELF). It is shown that highly efficient and robust SLF experiments can be designed based on double channel windowless homonuclear decoupling sequences. Compared to rotating frame techniques based on Hartmann-Hahn cross polarization, the new approach is less susceptible to the frequency offset and chemical shift interaction and can be applied in the structural studies of macromolecules that are uniformly labeled with isotopes such as (13)C and (15)N. Furthermore, isotropic mixing sequences allow for transfer of any magnetization component of one nucleus to the corresponding component of its dipolar coupled partner. The performance of HIMSELF is studied by analysis of the average Hamiltonian and numerical simulation and is experimentally demonstrated on a single crystalline sample of a dipeptide and a liquid crystalline sample exhibiting motionally averaged dipolar couplings.     

Measurement of carbon-proton dipolar couplings in liquid crystals using DAPT

Dipolar couplings provide valuable information on order and dynamics in liquid crystals. For measuring heteronuclear dipolar couplings in oriented systems, a new separated local field experiment is presented here. The method is based on the dipolar assisted polarization transfer (DAPT) pulse sequence proposed recently (Chem. Phys. Lett. 2007, 439, 407) for transfer of polarization between two spins I and S. DAPT utilizes the evolution of magnetization of the I and S spins under two blocks of phase shifted BLEW-12 pulses on the I spin separated by a 90 degree pulse on the S spin. Compared to the rotating frame techniques based on Hartmann-Hahn match, this approach is easy to implement and is independent of any matching conditions. DAPT can be utilized either as a proton encoded local field (PELF) technique or as a separated local field (SLF) technique, which means that the heteronuclear dipolar coupling can be obtained by following either the evolution of the abundant spin like proton (PELF) or that of the rare spin such as carbon (SLF). We have demonstrated the use of DAPT both as a PELF and as a SLF technique on an oriented liquid crystalline sample at room temperature and also have compared its performance with PISEMA. We have also incorporated modifications to the original DAPT pulse sequence for (i) improving its sensitivity and (ii) removing carrier offset dependence.     

1H to 2H uniform cross-polarization nuclear magnetic resonance using 2H Lee-Goldburg irradiation in static powders

Application of conventional cross polarization (CP) to (2)H results in only a narrowband enhancement of the powder line shape due to the quadrupole interaction. We propose a CP scheme to uniformly enhance (2)H spectra in static powders. In this method, a Hartmann-Hahn matched (2)H rf field is applied on the Lee-Goldburg (LG) condition to remove the zeroth-order quadrupole interaction. In order to achieve a uniformly enhanced (2)H powder line shape with a limited (2)H rf intensity, the (1)H rf amplitude in CP is stepwise altered during the contact time. We develop a spin-thermodynamic theory to describe polarization transfer due to CP with LG irradiation (LG-CP) under the influence of the quadrupole interaction, which can successfully reproduce the LG-CP line shapes observed under various experimental conditions. Experimental and simulated (2)H powder spectra are reported for some compounds.     

Thermodynamics of adiabatic cross polarization

Using a spin-temperature approach, we describe a scheme of adiabatic cross polarization, based on demagnetization/remagnetization, when the Zeeman order of abundant nuclei in the laboratory frame is first adiabatically converted into the dipolar order, and then, into the Zeeman order of rare nuclei. The scheme, implemented with two low-power frequency-sweeping pulses, is very efficient for static samples and can significantly increase polarization of rare nuclei, compared to the conventional Hartmann-Hahn cross polarization. The experimental examples are presented for a solid, liquid crystal, and small molecules in a liquid-crystalline solvent.     

Dual acquisition magic-angle spinning solid-state NMR-spectroscopy: simultaneous acquisition of multidimensional spectra of biomacromolecules

Fast data collection: a general method for dual data acquisition of multidimensional magic-angle spinning solid-state NMR experiments is presented. The method uses a simultaneous Hartmann-Hahn cross-polarization from (1)H to (13)C and (15)N nuclei and exploits the long-living (15)N polarization for parallel acquisition of two multidimensional experiments.     

Selective polarization transfer using a single rf field

NMR is a popular and mature technique used in fields as diverse as chemistry, biology, or material science. One reason for this versatility lies in its ability to correlate the nuclei that are present in one molecule to another. This provides the researcher with correlation maps allowing for studies of the molecules at an atomic level. Selective experiments allow isolation of one such correlation to focus on spins of interest. This leads to a savings in precious experimental time by reducing the dimension of the experiment, which in turn may enable one to record more elaborate experiments that would otherwise not be amenable within reasonable acquisition times. Here, we present an alternative method to selectively transfer magnetization using a single rf field. This technique, which we call single field polarization transfer, allows to obtain longitudinal two-spin order of two scalar-coupled spins when only one of them is irradiated. The method is easy to implement and does not depend on stringent conditions, such as Hartmann-Hahn matching for selective cross-polarization transfers or very long inversion pulses and identification of coupling satellites in selective population inversion experiments.     

Direct measurement of dynamic frequency shift induced by cross-correlations in 15N-enriched proteins.

We describe a new NMR experimental scheme that allows the direct determination of the dynamic frequency shift induced by chemical shift anisotropy/dipolar interaction (CSA/DD) cross-correlations in 15N-enriched proteins. Its principle consists of comparing two rates of polarisation transfer between the amide proton and nitrogen. The first rate, which is independent of the dynamic frequency shift, is based on a selective Hartmann-Hahn coherence transfer. The second rate, which depends on the dynamic frequency shift, is based on a free evolution of the transverse magnetisation. We report experimental validation of this approach by measuring the average dynamic frequency shift due to CSA/DD cross-correlations in the calcium-binding protein D9k. The method may also be applicable to the measurement of dynamic frequency shift induced by cross-correlations between the Curie spin and dipolar interactions.     

SHARP-TACSY: triple-band tailored correlated spectroscopy for base-to-sugar transfer in nucleic acid residues with intermediate time scale motions

Established experiments to identify the sugar-to-base connectivity in isotopically labeled RNA require long transfer periods and are inefficient for residues undergoing intermediate time scale motions (microsecond to millisecond). Here, an alternative transfer experiment is introduced, whereby the C1'-N1/9-C6/8 spin system is selectively brought to the so-called Hartmann-Hahn condition using selectiveheteronuclear planar triple-band tailored correlated spectroscopy (SHARP-TACSY). Results are shown for the fully labeled 30-mer oligonucleotide TAR RNA with particular attention placed on residues from and close to the bulge and the loop. For these residues, the faster relaxation can be attributed to exchange contributions stemming from transient stacking and unstacking of the bases and/or from the isomerization of the ribose sugar pucker. The new experiment shows improved signal-to-noise for residues exhibiting large microsecond-millisecond time scale motions with respect to established experiments, thus providing a valid alternative for resonance assignment in mobile RNA regions.     

Background-Free Detection of Spin-Exchange Dynamics at Ultra-Low Magnetic Field

Ultra-low field nuclear magnetic resonance spectroscopy (NMR) and imaging (MRI) inherently suffer from a low signal-to-noise ratio due to the small thermal polarization of nuclear spins. Transfer of polarization from a pre-polarized spin system to a thermally polarized spin system via the Spin Polarization Induced Nuclear Overhauser Effect (SPINOE) could potentially be used to overcome this limitation. SPINOE is particularly advantageous at ultra-low magnetic field, where the transferred polarization can be several orders of magnitude higher than thermal polarization. Here we demonstrate direct detection of polarization transfer from highly polarized ¹²⁹Xe gas spins to ¹H spins in solution via SPINOE. At ultra-low field, where thermal nuclear spin polarization is close to background noise levels and where different nuclei can be simultaneously detected in a single spectrum, the dynamics of the polarization transfer can be observed in real time. We show that by simply bubbling hyperpolarized ¹²⁹Xe into solution, we can enhance ¹H polarization levels by a factor of up to 151-fold. While our protocol leads to lower enhancements than those previously reported under extreme Xe gas pressures, the methodology is easily repeatable and allows for on-demand enhanced spectroscopy. SPINOE at ultra-low magnetic field could also be employed to study ¹²⁹Xe interactions in solutions.     

Coherent Evolution of Signal Amplification by Reversible Exchange in Two Alternating Fields (alt-SABRE)

Parahydrogen (pH₂) is a convenient and cost-efficient source of spin order to enhance the magnetic resonance signal. Previous work showed that transient interaction of pH₂ with a metal organic complex in a signal amplification by reversible exchange (SABRE) experiment enabled more than 10 % polarization for some ¹⁵N molecules. Here, we analyzed a variant of SABRE, consisting of a magnetic field alternating between a low field of ∼1 μT, where polarization transfer is expected to take place, and a higher field >50 μT (alt-SABRE). These magnetic fields affected the amplitude and frequency of polarization transfer. Deviation of a lower magnetic field from a “perfect” condition of level anti-crossing increases the frequency of polarization transfer that can be exploited for polarization of short-lived transient SABRE complexes. Moreover, the coherences responsible for polarization transfer at a lower field persisted during magnetic field variation and continued their spin evolution at higher field with a frequency of 2.5 kHz at 54 μT. The latter should be taken into consideration for an efficient alt-SABRE. Theoretical and experimental findings were exemplified with Iridium N-heterocyclic carbene SABRE complex and ¹⁵N-acetonitrole, where a 30 % higher ¹⁵N polarization with alt-SABRE compared to common SABRE was reached.     

Mismatched Hartmann-Hahn conditions cause proton-mediated intermolecular magnetization transfer between dilute low-spin nuclei in NMR of static solids

Mismatched Hartmann-Hahn conditions between the protons and dilute spins (such as 15N) are found to cause intermolecular magnetization transfer between the low-gamma nuclei over long distances. This transfer is purely proton mediated and occurs even in the absence of direct 15N-15N couplings. This has been demonstrated experimentally using a static single crystal of n-acetyl Leucine with intermolecular distances between the 15N nuclei exceeding 6.5 A. A quantum-mechanical explanation of this phenomenon is given based on the average-Hamiltonian theory which was confirmed by detailed numerical many-spin simulations. The theory and experiment presented in the present paper may help in the development of solid-state NMR methods for studying interhelical contacts in membrane proteins, as well as for their spectral assignment.     

Off-resonance R(1rho) NMR studies of exchange dynamics in proteins with low spin-lock fields: an application to a Fyn SH3 domain

An (15)N NMR R(1rho) relaxation experiment is presented for the measurement of millisecond time scale exchange processes in proteins. On- and off-resonance R(1rho) relaxation profiles are recorded one residue at a time using a series of one-dimensional experiments in concert with selective Hartmann-Hahn polarization transfers. The experiment can be performed using low spin-lock field strengths (values as low as 25 Hz have been tested), with excellent alignment of magnetization along the effective field achieved. Additionally, suppression of the effects of cross-correlated relaxation between dipolar and chemical shift anisotropy interactions and (1)H-(15)N scalar coupled evolution is straightforward to implement, independent of the strength of the (15)N spin-locking field. The methodology is applied to study the folding of a G48M mutant of the Fyn SH3 domain that has been characterized previously by CPMG dispersion experiments. It is demonstrated through experiment that off-resonance R(1rho) data measured at a single magnetic field and one or more spin-lock field strengths, with amplitudes on the order of the rate of exchange, allow a complete characterization of a two-site exchange process. This is possible even in the case of slow exchange on the NMR time scale, where complementary approaches involving CPMG-based experiments fail. Advantages of this methodology in relation to other approaches are described.     

Solid effect dynamic nuclear polarization and polarization pathways

Using dynamic nuclear polarization (DNP)/nuclear magnetic resonance instrumentation that utilizes a microwave cavity and a balanced rf circuit, we observe a solid effect DNP enhancement of 94 at 5 T and 80 K using trityl radical as the polarizing agent. Because the buildup rate of the solid effect increases with microwave field strength, we obtain a sensitivity gain of 128. The data suggest that higher microwave field strengths would lead to further improvements in sensitivity. In addition, the observation of microwave field dependent enhancements permits us to draw conclusions about the path that polarization takes during the DNP process. By measuring the time constant for the polarization buildup and enhancement as a function of the microwave field strength, we are able to compare models of polarization transfer, and show that the major contribution to the bulk polarization arises via direct transfer from electrons, rather than transferring first to nearby nuclei and then transferring to bulk nuclei in a slow diffusion step. In addition, the model predicts that nuclei near the electron receive polarization that can relax, decrease the electron polarization, and attenuate the DNP enhancement. The magnitude of this effect depends on the number of near nuclei participating in the polarization transfer, hence the size of the diffusion barrier, their T(1), and the transfer rate. Approaches to optimizing the DNP enhancement are discussed.     

Polarization transfer in CIDNP experiments via scalar paramagnetic exchange mechanisms

A theory is outlined explaining how chemically induced nuclear polarization can be transfered by a diamagnetic-paramagnetic exchange reaction from a group of nuclei to another group not originally polarized. The mechanism depends critically on the scalar nuclear spin-spin coupling in the diamagnetic state. Experiments on the photochemical charge transfer between N,N-diethylaniline and pyrene are given as an example of this polarization transfer.     

Signals in solid-state photochemically induced dynamic nuclear polarization recover faster than signals obtained with the longitudinal relaxation time

During the photocycle of quinone-blocked photosynthetic reaction centers (RCs), photochemically induced dynamic nuclear polarization (photo-CIDNP) is produced by polarization transfer from the initially totally electron polarized electron pair and can be observed by 13C magic-angle spinning (MAS) NMR as a strong modification of signal intensities. The same processes creating net nuclear polarization open up light-dependent channels for polarization loss. This leads to coherent and incoherent enhanced signal recovery, in addition to the recovery due to light-independent longitudinal relaxation. Coherent mixing between electron and nuclear spin states due to pseudosecular hyperfine coupling within the radical pair state provides such a coherent loss channel for nuclear polarization. Another polarization transfer mechanism called differential relaxation, which is based on the long lifetime of the triplet state of the donor, provides an efficient incoherent relaxation path. In RCs of the purple bacterium Rhodobacter sphaeroides R26, the photochemical active channels allow for accelerated signal scanning by a factor of 5. Hence, photo-CIDNP MAS NMR provides the possibility to drive the NMR technique beyond the T1 limit.     

Theoretical study of dynamic electron-spin-polarization via the doublet-quartet quantum-mixed state and time-resolved ESR spectra of the quartet high-spin state

The mechanism of the unique dynamic electron polarization of the quartet (S = 3/2) high-spin state via a doublet-quartet quantum-mixed state and detail theoretical calculations of the population transfer are reported. By the photo-induced electron transfer, the quantum-mixed charge-separate state is generated in acceptor-donor-radical triad (A-D-R). This mechanism explains well the unique dynamic electron polarization of the quartet state of A-D-R. The generation of the selectively populated quantum-mixed state and its transfer to the strongly coupled pure quartet and doublet states have been treated both by a perturbation approach and by exact numerical calculations. The analytical solutions show that generation of the quantum-mixed states with the selective populations after de-coherence and/or accompanying the (complete) dephasing during the charge-recombination are essential for the unique dynamic electron polarization. Thus, the elimination of the quantum coherence (loss of the quantum information) is the key process for the population transfer from the quantum-mixed state to the quartet state. The generation of high-field polarization on the strongly coupled quartet state by the charge-recombination process can be explained by a polarization transfer from the quantum-mixed charge-separate state. Typical time-resolved ESR patterns of the quantum-mixed state and of the strongly coupled quartet state are simulated based on the generation mechanism of the dynamic electron polarization. The dependence of the spectral pattern of the quartet high-spin state has been clarified for the fine-structure tensor and the exchange interaction of the quantum-mixed state. The spectral pattern of the quartet state is not sensitive towards the fine-structure tensor of the quantum-mixed state, because this tensor contributes only as a perturbation in the population transfer to the spin-sublevels of the quartet state. Based on the stochastic Liouville equation, it is also discussed why the selective population in the quantum-mixed state is generated for the "finite field" spin-sublevels. The numerical calculations of the elimination of the quantum coherence (de-coherence and/or dephasing) are demonstrated. A new possibility of the enhanced intersystem crossing pathway in solution is also proposed.     

Electron Transfer in Frozen Media

Classical theories of electron transfer are modified to take into account the differences between electron transfer in a rigid medium and in a fluid. Intramolecular vibrations and part of the dielectric polarization are assumed to remain dynamic in rigid media while the remaining part of the polarization, arising from dipole reorientations, is frozen. In rigid media, electron transfer occurs with the solvent locked into the dipole orientations of the initial state. This causes an increase in the free energy change and a decrease in the solvent reorganizational energy. It also increases the activation free energy for electron transfer. For photoinduced electron transfer, the analysis is more complex because multiple states are involved. The activation free energy can either be greater or less than in a fluid depending on charge distributions before and after electron transfer. The same analysis can be applied to interconversion between excited states in rigid media.