Journal of Magnetic Resonance Open最新文献

A rigorous quantification of the thermodynamics, kinetic and structural changes can be approached by relaxation dispersion methods, when such motions are in the microsecond to millisecond timescale. Albeit some past efforts, it is still unclear how cosolutes modulate the fitted parameters extracted from relaxation dispersion analyses. Here, we have studied how the systematic measurement of ¹⁵N relaxation dispersion on a well-studied enzyme, undergoing two-state chemical exchange, are affected in terms of the populations of the minor state (thermodynamics) and the exchange rates (kinetics) in the presence of different cosolutes, from ions to denaturants. The cosolute-induced changes are modest but can significantly affect protein function. Specifically, exchange rates can be associated to a subtle leverage in the exchange rates. For most canonical residues, i.e. without spurious effects caused by the cosolute, the chemical shift difference ($\Delta \omega$) between both states is essentially unaffected with respect to the expected shift of water resonance. Yet, solvent exposed residues do not always follow this canonical trend. Instead, residue-cosolute interactions are large enough to induce a residue specific shift that ultimately modulates the fitted parameter $\Delta \omega$. Unfortunately, solvent accessibility analysis is not accurate enough to a priori discriminate between canonical and non-canonical residues and a full experimental analysis at varying concentrations of the cosolute is required. This observation aims to act as a word of caution for measurements of relaxation dispersion at a single-solvent condition that contains large amounts of cosolutes.

Human blood is the most widely used biospecimen in the clinic and the metabolomics field. While both mass spectrometry and NMR spectroscopy are the two premier analytical platforms in the metabolomics field, NMR exhibits several unsurpassed characteristics for blood metabolite analysis, the most important of which are its ability to identify unknown metabolites and its quantitative nature. However, the relatively small number of metabolites accessible by NMR has restricted the scope of its applications. Enhancing the limit of identified metabolites in blood will therefore greatly impact NMR-based metabolomics. Continuing our efforts to address this major issue, our current study describes the identification of 12 metabolites, which expands the number of quantifiable blood metabolites by ∼15%. These results, in combination with our earlier efforts, now provide access to nearly 90 metabolites, which is the highest to date for a simple 1D ¹H NMR experiment that is widely used in the metabolomics field. Metabolites were identified based on the comprehensive investigation of human blood and plasma using 1D/2D NMR techniques. The newly identified metabolites were validated based on chemical shift databases, spectra of authentic compounds obtained under conditions identical to blood/plasma, and, finally, spiking experiments using authentic compounds. Considering the high reproducibility of NMR and the sensitivity of chemical shifts to altered sample conditions, experimental protocols and peak annotations are provided for the newly identified metabolites, which serve as a template for identification of blood metabolites for routine applications. Separately, the identified metabolites were evaluated for their sensitivity to preanalytical conditions. The results reveal that among the newly identified metabolites, inosine monophosphate (IMP) and nicotinamide are associated with labile coenzymes and their levels are sensitive to preanalytical conditions. The study demonstrates the expansion of quantifiable blood metabolites using NMR to a new height and is expected to greatly impact blood metabolomics.

Nuclear Magnetic Resonance (NMR) has been widely used in Petroleum Science and Engineering to study geological formations (porous media) in laboratories or under well-logging conditions. In both cases, NMR is still evolving to provide more accurate data on well productivity. In well-logging, NMR is one of the main tools used in determining the economic viability of an oil well due to the reliability of measurements of fluid types, porosity, pore size, and permeability of the reservoir under analysis. There are two kinds of NMR well-logging techniques: Wireline and Logging While Drilling (LWD). In the latter, due to the drilling process, measurements are made with the NMR tool translating, vibrating and, in some cases, rotating relatively to the geological formation. To understand the behavior of NMR signals measured under LWD conditions, not yet including displacement and drill vibration, we have recently developed a single-sided magnet, probes, and a mechanical system that emulates a relative sinusoidal motion between the sample and the applied magnetic field. This equipment was used to emulate a LWD tool operating under normal pressure and temperature conditions.

Intrinsically disordered regions (IDRs) lack stable tertiary structure and instead rapidly interconvert between different conformations. This structural plasticity enables IDRs to act as key players in cellular signaling pathways. Transcription factors are enriched in IDRs, many of which are stabilized by or acquire tertiary structure in the presence of DNA or other binding partners. Using the T-cell factor/lymphoid enhancer binding factor 1 (TCF/LEF-1) transcription factor as a model system, we characterized the structure and dynamics of the high-mobility group (HMG) domain in the absence of DNA. Inclusion of the IDRs that flank the HMG domain led to enhanced solubility of the protein. Secondary ¹³Cα chemical shifts, ¹H nuclear Overhauser effects, ¹⁵N spin relaxation, and ¹H^N solvent paramagnetic relaxation enhancements indicate that the three helices in the HMG domain are oriented similarly to the DNA-bound form of the protein. By contrast, the flanking IDRs do not show evidence of structure. Helix 1 and helix 3 appear to be less stable in the DNA-free conformation, indicating some form of conformational exchange or local motion in the absence of DNA. Given the high degree of sequence conservation in the TCF/LEF family of transcription factors, our results should apply to other members of the family.

This primer covers de-bugging NMR hardware, at the level of major components (not the circuit board level). It is intended for users of NMR who are unlikely to build their own spectrometer, but will need to localize the malfunction to individual small sub-systems. Brand names and some part numbers of appropriate test equipment and lab-gadgets are given. But many users will find themselves in need of special purpose NMR probes, so construction of a dedicated probe is not beyond consideration. Thus, simple single-resonance probe circuits are discussed at greater length. The need for tuning and coupling of the probe tuned circuit is treated as an exercise in impedance matching.

A transceiver volume coil for High Field Magnetic Resonance Imaging was developed based on Temnikov’s NMR probe coil. The critical advantage of this design is that requires only one trimmer capacitor for both tuning and matching. Electromagnetic simulations of our coil were compared to a four-rung birdcage coil to evaluate their performances. A transceiver coil prototype, called the double-cross coil and a birdcage coil were built to compare and contrast their feasibility. Both coil prototypes were operated in the linear mode. Experimental quality factors were $Q_u/Q_l=101/45$ for the double-cross coil and $Q_u/Q_l=105/43$ for the birdcage coil. Phantom images were acquired to test the viability of each coil for high field magnetic resonance imaging applications. The signal-to-noise ratios, noise factors and uniformity values were also calculated: $85.26/0.74/44.48$ for the double-cross coil and $76.54/1.9/65.65$ for the birdcage coil. These volume coils showed similar performance, but the double-cross coil has better uniformity and a slightly lower noise figure than the birdcage coil. A remarkable agreement of experimental and simulated $B_1$ was obtained for the double-cross coil. These experimental and numerical results demonstrate the feasibility of the double-cross volume coil with better uniformity for rodent magnetic resonance imaging at 7 T.

Serial acquisition of one-dimensional NMR spectra appears in many contexts, e.g. in variable-temperature studies or reaction monitoring. In a conventional approach, the spectra are processed separately, and standard peak-picking is performed in each of them. Yet, when chemical shifts change linearly between spectra, the Radon transform (RT) is more effective than conventional data processing, since it provides sensitivity and resolution gains. RT results in a two-dimensional (2D) spectrum with one dimension corresponding to resonance frequencies and the other to their rates of change. However, the lineshapes in 2D RT spectra are not 2D lorentzians, and thus available spectral peak-pickers cannot effectively deal with them. We propose a solution to this problem — a peak-picker dedicated to 2D RT spectra and based on a U-Net neural network. The software contains a user-friendly graphical interface. We test the program on three challenging serial data sets to demonstrate the robustness to peak overlap, complex multiplet structures and low signal-to-noise ratio.

The static wideline and high-resolution magic angle spinning (MAS) ²⁵Mg NMR lineshapes measured for isotopically enriched enstatite and diopside glasses indicate a wide distribution of the electric-field gradient (EFG) components at the ²⁵Mg position caused by disorder. The correct characterization of this distribution requires simulations with special attention paid to the experimental parameters. Here, both the static spectra obtained by wideband excitation and the MAS-NMR spectra obtained via rotor-synchronized Hahn spin echo acquisition were successfully simulated with the Czjzek distribution model, using one consistent set of parameters. Average nuclear electric quadrupole coupling constants near 8 MHz and a distribution width around 4 MHz were obtained for both materials, which suggests that earlier results on these glasses need to be re-examined. The results of this study outline a general strategy for characterizing the local environments of strongly coupled quadrupolar nuclei in amorphous and glassy materials. Despite the discrepancies between the interaction parameters extracted from our data and those published in earlier NMR work, the best fit to the data indicate an average isotropic chemical shift near 13 ppm (vs. aqueous MgCl₂ solution) for both the enstatite and diopside glasses. Assuming the applicability of the current database relating the ²⁵Mg chemical shifts with coordination numbers in crystalline compounds, this value suggests that the Mg²⁺ ions are six-coordinated. This conclusion, however, is based on the simplifying assumption that the ²⁵Mg spectrum comprises a Czjzek distribution centered at a single isotropic chemical shift value and stands in contrast to the results of recent isotope-selective neutron diffraction studies which give an average Mg-O coordination number of 4.4–4.5 for both glasses. However, reasonable fits to the MAS-NMR spectra can also be obtained when constraining the average isotropic chemical shift to 50 ppm, a value typical of four-coordinated Mg. We conclude that multiple Mg sites with different coordination numbers may well be present and that, in the present glasses, ²⁵Mg NMR at typically used spinning rates and magnetic field strengths (20 kHz, 14.1 T) is not capable of resolving them due to excessive broadening by quadrupolar interactions.

Gadolinium-based MRI contrast agent Dotarem is commonly used in the clinic. Free Gd³⁺ is very toxic. Because the crystallographic radius of Gd³⁺ and Ca²⁺ is very similar, Gd³⁺ can compete with Ca²⁺ in the regulations of cellular functions. Gadolinium is retained in organs and tissues, but the effects of gadolinium retention are mostly unknown. There is also no information how gadolinium retention impacts immune cells and signaling pathways regulated by Ca²⁺ such as RhoA/mTORC1 and mTORC2, which control various, including actin cytoskeleton-dependent, cellular functions. We developed the 7T MRI-based method to analyze gadolinium-based MRI contrast agent retention in isolated immune cells (macrophages) and study the effects of gadolinium retention on the expression of RhoA, mTORC1, mTORC2, and mitochondria, Golgi, and ER markers. We showed that macrophages retain gadolinium for at least seven days after exposure. Gadolinium retention downregulated the expression of RhoA, mTORC1 component Raptor and mTORC2 component Rictor proteins, and dysregulated the expression level of organelle markers. The method described here can be used for monitoring gadolinium levels and effects in isolated cells, such as the immune cells in the blood of patients exposed to contrast-enhanced MRI.

Mutations of the breast cancer susceptibility 1 (BRCA1) gene are associated with high risks of breast and ovarian cancer, not least due to the vital role of the BRCA1 gene product in DNA double-strand repair. Strikingly, little is known about the structural dynamics of BRCA1-DNA complexes, despite their importance for maintaining a healthy cell cycle and the potential boost for rational targeting of BRCA1 by their detailed understanding. Aiming to address this shortcoming, we present a model resulting from the direct binding of the intrinsically disordered region (IDR) of BRCA1 to DNA oligomers. To this end, we developed a workflow combining in-silico structural predictions, computational docking, and molecular dynamics simulations with chemical shift perturbations in ¹H-³¹P crosspeaks obtained by nuclear magnetic resonance spectroscopy of a binding DNA oligomer. Our data show that the BRCA1-DNA complexes are stabilized, mainly through ‘head-on’ interaction between BRCA1 and the DNA double-strand ends. ‘Side-on’ binding to the DNA major grove was insufficient to form stable complexes. When bound to the nucleic acid, the IDR maintained high degrees of flexibility in our simulations reminiscent of ‘fuzzy’ complexes. Illustrating the structural dynamics underlying BRCA1-DNA complexes is essential for the bottom-up reconstruction of the role BRCA1 plays in DNA double-strand break repair. The presented work makes a step in this direction, aiming to complement existing assays with models that can assist in the functional screening of hereditary breast and ovarian cancer (HBOC)-relevant mutations.