A matter of minutes: Dynamic nuclear polarization (DNP) is applied to enhance the signal of solid‐state NMR spectra of metal–organic framework (MOF) materials. The signal enhancement enables the ...acquisition of high‐quality 1D 13C solid‐state NMR spectra, 2D 1H–13C dipolar HETCOR and 1D 15N solid‐state NMR spectra with natural isotopic abundance in experiment times on the order of minutes. MW=microwaves.
Dynamic Nuclear Polarization (DNP) experiments transfer polarization from electron spins to nuclear spins with microwave irradiation of the electron spins for enhanced sensitivity in nuclear magnetic ...resonance (NMR) spectroscopy. Design and testing of a spectrometer for magic angle spinning (MAS) DNP experiments at 263 GHz microwave frequency, 400 MHz (1)H frequency is described. Microwaves are generated by a novel continuous-wave gyrotron, transmitted to the NMR probe via a transmission line, and irradiated on a 3.2 mm rotor for MAS DNP experiments. DNP signal enhancements of up to 80 have been measured at 95 K on urea and proline in water-glycerol with the biradical polarizing agent TOTAPOL. We characterize the experimental parameters affecting the DNP efficiency: the magnetic field dependence, temperature dependence and polarization build-up times, microwave power dependence, sample heating effects, and spinning frequency dependence of the DNP signal enhancement. Stable system operation, including DNP performance, is also demonstrated over a 36 h period.
A new nitroxide-based biradical having a long electron spin–lattice relaxation time (T 1e) has been developed as an exogenous polarization source for DNP solid-state NMR experiments. The performance ...of this new biradical is demonstrated on hybrid silica-based mesostructured materials impregnated with 1,1,2,2-tetrachloroethane radical containing solutions, as well as in frozen bulk solutions, yielding DNP enhancement factors (ε) of over 100 at a magnetic field of 9.4 T and sample temperatures of ∼100 K. The effects of radical concentration on the DNP enhancement factors and on the overall sensitivity enhancements (Σ†) are reported. The relatively high DNP efficiency of the biradical is attributed to an increased T 1e, which enables more effective saturation of the electron resonance. This new biradical is shown to outperform the polarizing agents used so far in DNP surface-enhanced NMR spectroscopy of materials, yielding a 113-fold increase in overall sensitivity for silicon-29 CPMAS spectra as compared to conventional NMR experiments at room temperature. This results in a reduction in experimental times by a factor >12 700, making the acquisition of 13C and 15N one- and two-dimensional NMR spectra at natural isotopic abundance rapid (hours). It has been used here to monitor a series of chemical reactions carried out on the surface functionalities of a hybrid organic–silica material.
Dynamic nuclear polarization (DNP) has become a powerful method to enhance spectroscopic sensitivity in the context of magnetic resonance imaging and nuclear magnetic resonance spectroscopy. We show ...that, compared to DNP at lower field (400 MHz/263 GHz), high field DNP (800 MHz/527 GHz) can significantly enhance spectral resolution and allows exploitation of the paramagnetic relaxation properties of DNP polarizing agents as direct structural probes under magic angle spinning conditions. Applied to a membrane-embedded K
+
channel, this approach allowed us to refine the membrane-embedded channel structure and revealed conformational substates that are present during two different stages of the channel gating cycle. High-field DNP thus offers atomic insight into the role of molecular plasticity during the course of biomolecular function in a complex cellular environment.
Comprehensive multiphase (CMP) NMR is a novel technology that integrates all the hardware from solution-, gel- and solid-state into a single NMR probe, permitting all phases to be studied in intact ...samples. Here comprehensive multiphase (CMP) NMR is used to study all components in a living organism for the first time. This work describes 4 new scientific accomplishments summarized as: (1) CMP-NMR is applied to a living animal, (2) an effective method to deliver oxygen to the organisms is described which permits longer studies essential for in-depth NMR analysis in general, (3) a range of spectral editing approaches are applied to fully differentiate the various phases solutions (metabolites) through to solids (shell) (4)
C isotopic labelling and multidimensional NMR are combined to provide detailed assignment of metabolites and structural components
. While not explicitly studied here the multiphase capabilities of the technique offer future possibilities to study kinetic transfer between phases (
nutrient assimilation, contaminant sequestration), molecular binding at interfaces (
drug or contaminant binding) and bonding across and between phases (
muscle to bone)
. Future work will need to focus on decreasing the spinning speed to reduce organism stress during analysis.
A peek inside: Dynamic nuclear polarization (DNP) enhances the spectroscopic sensitivity of solid‐state NMR measurements of uniformly (13C,15N)‐labeled preparations of Escherichia coli cells by more ...than an order of magnitude (see picture; MW=microwaves, ε=enhancement factor). The major molecular components in the cells can be characterized in this way.
We describe magic-angle spinning NMR experiments designed to elucidate the interstrand architecture of amyloid fibrils. Three methods are introduced for this purpose, two being based on the analysis ...of long-range 13C–13C correlation spectra and the third based on the identification of intermolecular interactions in 13C–15N spectra. We show, in studies of fibrils formed by the 86-residue SH3 domain of PI3 kinase (PI3-SH3 or PI3K-SH3), that efficient 13C–13C correlation spectra display a resonance degeneracy that establishes a parallel, in-register alignment of the proteins in the amyloid fibrils. In addition, this degeneracy can be circumvented to yield direct intermolecular constraints. The 13C–13C experiments are corroborated by 15N–13C correlation spectra obtained from a mixed 15N,12C/14N,13C sample which directly quantify interstrand distances. Furthermore, when the spectra are recorded with signal enhancement provided by dynamic nuclear polarization (DNP) at 100 K, we demonstrate a dramatic increase (from 23 to 52) in the number of intermolecular 15N–13C constraints detectable in the spectra. The increase in the information content is due to the enhanced signal intensities and to the fact that dynamic processes, leading to spectral intensity losses, are quenched at low temperatures. Thus, acquisition of low temperature spectra addresses a problem that is frequently encountered in MAS spectra of proteins. In total, the experiments provide 111 intermolecular 13C–13C and 15N–13C constraints that establish that the PI3-SH3 protein strands are aligned in a parallel, in-register arrangement within the amyloid fibril.
Since the isolation of soil organic matter in 1786, tens of thousands of publications have searched for its structure. Nuclear magnetic resonance (NMR) spectroscopy has played a critical role in ...defining soil organic matter but traditional approaches remove key information such as the distribution of components at the soil–water interface and conformational information. Here a novel form of NMR with capabilities to study all physical phases termed Comprehensive Multiphase NMR, is applied to analyze soil in its natural swollen-state. The key structural components in soil organic matter are identified to be largely composed of macromolecular inputs from degrading biomass. Polar lipid heads and carbohydrates dominate the soil–water interface while lignin and microbes are arranged in a more hydrophobic interior. Lignin domains cannot be penetrated by aqueous solvents even at extreme pH indicating they are the most hydrophobic environment in soil and are ideal for sequestering hydrophobic contaminants. Here, for the first time, a complete range of physical states of a whole soil can be studied. This provides a more detailed understanding of soil organic matter at the molecular level itself key to develop the most efficient soil remediation and agricultural techniques, and better predict carbon sequestration and climate change.
Dynamic nuclear polarization (DNP) utilizes the inherently larger polarization of electrons to enhance the sensitivity of conventional solid-state NMR experiments at low temperature. Recent advances ...in instrumentation development and sample preparation have transformed this field and have opened up new opportunities for its application to biological systems. Here, we present DNP-enhanced (13)C-(13)C and (15)N-(13)C correlation experiments on GNNQQNY nanocrystals and amyloid fibrils acquired at 9.4 T and 100 K and demonstrate that DNP can be used to obtain assignments and site-specific structural information very efficiently. We investigate the influence of temperature on the resolution, molecular conformation, structural integrity and dynamics in these two systems. In addition, we assess the low-temperature performance of two commonly used solid-state NMR experiments, proton-driven spin diffusion (PDSD) and transferred echo double resonance (TEDOR), and discuss their potential as tools for measurement of structurally relevant distances at low temperature in combination with DNP.
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