Three-dimensional Nuclear Magnetic Resonance Research Articles

The Use of Spectroscopic Methods to Study Organic Matter in Virgin and Arable Soils: A Scoping Review

The use of modern spectroscopic methods of analysis, which provide extensive information on the chemical nature of substances, significantly expands our understanding of the molecular composition and properties of soil organic matter (SOM) and its transformation and stabilization processes in various ecosystems and geochemical conditions. The aim of this review is to identify and analyze studies related to the application of nuclear magnetic resonance (NMR) and electron paramagnetic resonance (EPR) spectroscopy techniques to study the molecular composition and transformation of organic matter in virgin and arable soils. This article is mainly based on three research questions: (1) Which NMR spectroscopy techniques are used to study SOM, and what are their disadvantages and advantages? (2) How is the NMR spectroscopy technique used to study the molecular structure of different pools of SOM? (3) How is ESR spectroscopy used in SOM chemistry, and what are its advantages and limitations? Relevant studies published between 1996 and 2024 were searched in four databases: eLIBRARY, MDPI, ScienceDirect and Springer. We excluded non-English-language articles, review articles, non-peer-reviewed articles and other non-article publications, as well as publications that were not available according to the search protocols. Exclusion criteria for articles were studies that used NMR and EPR techniques to study non-SOM and where these techniques were not the primary methods. Our scoping review found that both solid-state and solution-state NMR spectroscopy are commonly used to study the structure of soil organic matter (SOM). Solution-phase NMR is particularly useful for studying soluble SOM components of a low molecular weight, whereas solid-phase NMR offers advantages such as higher 13C atom concentration for stronger signals and faster analysis time. However, solution-phase NMR has limitations including sample insolubility, potential signal aggregation and reduced sensitivity and resolution. Solid-state NMR is better at detecting non-protonated carbon atoms and identifying heterogeneous regions within structures. EPR spectroscopy, on the other hand, offers significant advantages in experimental biochemistry due to its high sensitivity and ability to provide detailed information about substances containing free radicals (FRs), aiding in the assessment of their reactivity and transformations. Understanding the FR structure in biopolymers can help to study the formation and transformation of SOM. The integration of two- and three-dimensional NMR spectroscopy with other analytical methods, such as chromatography, mass spectrometry, etc., provides a more comprehensive approach to deciphering the complex composition of SOM than one-dimensional techniques alone.

Unraveling Molecular Recognition of Glycan Ligands by Siglec-9 via NMR Spectroscopy and Molecular Dynamics Modeling.

Human sialic-acid-binding immunoglobulin-like lectin-9 (Siglec-9) is a glycoimmune checkpoint receptor expressed on several immune cells. Binding of Siglec-9 to sialic acid containing glycans (sialoglycans) is well documented to modulate its functions as an inhibitory receptor. Here, we first assigned the amino acid backbone of the Siglec-9 V-set domain (Siglec-9d1), using well-established triple resonance three-dimensional nuclear magnetic resonance (NMR) methods. Then, we combined solution NMR and molecular dynamic simulation methods to decipher the molecular details of the interaction of Siglec-9 with the natural ligands α2,3 and α2,6 sialyl lactosamines (SLN), sialyl Lewis X (sLeX), and 6-O sulfated sLeX and with two synthetically modified sialoglycans that bind with high affinity. As expected, Neu5Ac is accommodated between the F and G β-strands at the canonical sialic acid binding site. Addition of a heteroaromatic scaffold 9N-5-(2-methylthiazol-4-yl)thiophene sulfonamide (MTTS) at the C9 position of Neu5Ac generates new interactions with the hydrophobic residues located at the G-G' loop and the N-terminal region of Siglec-9. Similarly, the addition of the aromatic substituent (5-N-(1-benzhydryl-1H-1,2,3-triazol-4-yl)methyl (BTC)) at the C5 position of Neu5Ac stabilizes the conformation of the long and flexible B'-C loop present in Siglec-9. These results expose the underlying mechanism responsible for the enhanced affinity and specificity for Siglec-9 for these two modified sialoglycans and sheds light on the rational design of the next generation of modified sialoglycans targeting Siglec-9.

Solid-state NMR MAS CryoProbe enables structural studies of human blood protein vitronectin bound to hydroxyapatite

The low sensitivity of nuclear magnetic resonance (NMR) is a major bottleneck for studying biomolecular structures of complex biomolecular assemblies. Cryogenically cooled probe technology overcomes the sensitivity limitations enabling NMR applications to challenging biomolecular systems. Here we describe solid-state NMR studies of the human blood protein vitronectin (Vn) bound to hydroxyapatite (HAP), the mineralized form of calcium phosphate, using a CryoProbe designed for magic angle spinning (MAS) experiments. Vn is a major blood protein that regulates many different physiological and pathological processes. The high sensitivity of the CryoProbe enabled us to acquire three-dimensional solid-state NMR spectra for sequential assignment and characterization of site-specific water-protein interactions that provide initial insights into the organization of the Vn-HAP complex. Vn associates with HAP in various pathological settings, including macular degeneration eyes and Alzheimer's disease brains. The ability to probe these assemblies at atomic detail paves the way for understanding their formation.

Amyloid fibril structures of tau: Conformational plasticity of the second microtubule-binding repeat.

The intrinsically disordered protein tau associates with microtubules in neurons but aggregates into cross-β amyloid fibrils that propagate in neurodegenerative brains. Different tauopathies have different structures for the rigid fibril core. To understand the molecular basis of tau assembly into different polymorphs, here we use solid-state nuclear magnetic resonance (NMR) spectroscopy to determine the structure of a tau protein that includes all microtubule-binding repeats and a proline-rich domain. This P2R tau assembles into well-ordered filaments when induced by heparin. Two- and three-dimensional NMR spectra indicate that R2 and R3 repeats constitute the rigid β-sheet core of the fibril. Unexpectedly, the amino-terminal half of R2 forms a β-arch at ambient temperature (24°C) but a continuous β-strand at 12°C, which dimerizes with the R2 of another protofilament. This temperature-dependent structure indicates that R2 is conformationally more plastic than the R3 domain. The distinct conformational stabilities of different microtubule-binding repeats give insight into the energy landscape of tau fibril formation.

Review of Recent Developments in NMR Core Analysis

The last 10 years have witnessed numerous new developments and advancements in nuclear magnetic resonance (NMR) rock core analysis both in conventional and unconventional reservoirs. On the hardware side, NMR instruments are now capable of handling hundreds of thousands of radio frequency (RF) pulses, and the pulses can be more closely spaced than ever. This allows measuring samples with very fast relaxation times, which was not possible before. Higher field NMR spectrometers operating at frequencies of ~12 MHz and ~20 MHz (in addition to the standard low-field ~2 MHz instruments) were introduced, which drastically improved the signal-to-noise ratio (SNR) and even allowed two-dimensional (2D) and three-dimensional (3D) NMR imaging to be performed on the samples to complement traditional relaxation measurements. Another consequence of using higher field instruments is that even samples with a very low amount of NMR signal can now provide useful data in a reasonable amount of time. Additional hardware improvements include the development of NMR-compatible high-temperature and high-pressure vessels and faster and more robust electronics and computers, which allow faster data acquisition, processing, and a more user-friendly environment for new NMR pulse sequence design and implementation. Following the hardware improvements, new applications were developed for rock core analysis, including modified Carl-Purcell-Meiboom-Gill (CPMG) pulse sequences for minimizing sample heating. NMR-based wettability, relative permeability, capillary pressure, cuttings, and long core measurements have also been successful. In unconventionals, NMR-based gas isotherm measurements and fluid-typing measurements are now common practices. This review paper sets out to summarize the recent NMR developments in the industry as well as provide references where more details can be found on each topic.

High-field magnetic resonance imaging: Challenges, advantages, and opportunities for novel contrast agents

There is a need for non-invasive diagnostic tools to detect and monitor the occurrence of diseases. Ideally, this can be done without resorting to ionizing radiation, especially when multiple rounds of imaging are required. Magnetic resonance imaging (MRI), a form of three-dimensional nuclear magnetic resonance, has become a common tool of choice for diagnosticians. Due to the low contrast difference between healthy and diseased tissue, contrast agents—magnetic species administered to the patient prior to imaging—are routinely used for contrast improvement. High-field (B0 ≥ 4.7 T, 1H Larmor frequency ≥ 200 MHz) MRI offers advantages in terms of better signal-to-noise ratio, as well as improved spectral resolution for certain applications. New contrast agents are being developed for high-field MRI, the topic of this review. After discussing the purpose of contrast agents and the advantages and potential issues of high-field MRI, we discuss recent developments in the field of contrast agent design, synthesis, and applications, citing examples of high-field MRI-ready molecular contrast agents, as well as nanoparticulate contrast agents based on various inorganic materials (e.g., coordination polymers, transition metal oxides, or lanthanide halides). We will discuss how certain aspects (composition, shape, ligands) affect the contrasting abilities of these agents. Finally, we highlight recent developments in the promising field of multifunctional probes, wherein multiple imaging and/or therapeutic modalities are combined in a single species. As high-field MRI becomes more commonplace in the clinical setting, such new contrast agents are needed to provide optimized imaging. This will facilitate the clinician's task in resolving pathologies for more efficient diagnosis and patient treatment.

Constraints on the Structure of Fibrils Formed by a Racemic Mixture of Amyloid-β Peptides from Solid-State NMR, Electron Microscopy, and Theory.

Previous studies have shown that racemic mixtures of 40- and 42-residue amyloid-β peptides (d,l-Aβ40 and d,l-Aβ42) form amyloid fibrils with accelerated kinetics and enhanced stability relative to their homochiral counterparts (l-Aβ40 and l-Aβ42), suggesting a "chiral inactivation" approach to abrogating the neurotoxicity of Aβ oligomers (Aβ-CI). Here we report a structural study of d,l-Aβ40 fibrils, using electron microscopy, solid-state nuclear magnetic resonance (NMR), and density functional theory (DFT) calculations. Two- and three-dimensional solid-state NMR spectra indicate molecular conformations in d,l-Aβ40 fibrils that resemble those in known l-Aβ40 fibril structures. However, quantitative measurements of 13C-13C and 15N-13C distances in selectively labeled d,l-Aβ40 fibril samples indicate a qualitatively different supramolecular structure. While cross-β structures in mature l-Aβ40 fibrils are comprised of in-register, parallel β-sheets, our data indicate antiparallel β-sheets in d,l-Aβ40 fibrils, with alternation of d and l molecules along the fibril growth direction, i.e., antiparallel "rippled sheet" structures. The solid-state NMR data suggest the coexistence of d,l-Aβ40 fibril polymorphs with three different registries of intermolecular hydrogen bonds within the antiparallel rippled sheets. DFT calculations support an energetic preference for antiparallel alignments of the β-strand segments identified by solid-state NMR. These results provide insight into the structural basis for Aβ-CI and establish the importance of rippled sheets in self-assembly of full-length, naturally occurring amyloidogenic peptides.

Three-dimensional nuclear magnetic resonance spectroscopy: a complementary tool to multiparametric magnetic resonance imaging in the identification of aggressive prostate cancer at 3.0T.

The limitations of the assessment of tumor aggressiveness by Prostate Imaging Reporting and Data System (PI-RADS) and biopsies suggest that the diagnostic algorithm could be improved by quantitative measurements in some chosen indications. We assessed the tumor high-risk predictive performance of 3.0 Tesla (3.0T) multiparametric magnetic resonance imaging (mp-MRI) combined with nuclear magnetic resonance spectroscopic sequences (NMR-S) in order to show that the metabolic analysis could bring out an evocative result for the aggressive form of prostate cancer. We conducted a retrospective study of 26 patients (mean age, 62.4 years) who had surgery for prostate cancer between 2009 and 2016 after pre-therapeutic assessment with 3.0T mp-MRI and NMR-S. Groups within the intermediate range of the D'Amico risk classification were divided into two categories, low risk (n=20) and high risk (n=6), according to the International Society of Urological Pathology (ISUP) 2-3 limit. Histoprognostic discordances within various risk groups were compared with the corresponding predictive MRI values. The performance of predictive models was assessed based on sensitivity, specificity, and the area under the curve (AUC) of receiver operating characteristic (ROC) curves. After prostatectomy, histological analysis reclassified 18 patients as high-risk, including 16 who were T3 MRI grade, of whom 13 (81.3%) were found to be pT3. Among the patients who had cT1 or cT2 digital rectal examinations, the T3 MRI factor multiplied by 8.7 [odds ratio (OR), 8.7; 95% confidence interval (CI), 1.3-56.2; P=0.024] the relative risk of being pT3 and by 5.8 (OR, 5.8; 95% CI, 0.95-35.7; P=0.05) the relative risk of being pGleason (pGS) > GS-prostate biopsy. Spectroscopic data showed that the choline concentration was significantly higher (P=0.001) in aggressive disease. The predictive model of tumor aggressiveness combining mp-MRI plus NMR-S was better than the mp-MRI model alone (AUC, 0.95 vs. 0.86). Information obtained by mp-MRI coupled with spectroscopy may improve the detection of occult aggressive disease, helping in the discrimination of intermediate risks.

Pore-fluid characterizations and microscopic mechanisms of sedimentary rocks with three-dimensional NMR: Tight sandstone as an example

Pore-fluid is directly related to the reservoir quality. Three-dimensional (3D) nuclear magnetic resonance (NMR) can be used for charactering pore-fluid due to its sensitivity to fluid components saturated in sedimentary rocks. In this paper, taken a tight sandstone as an example, investigating pore-fluid characterizations and microscopic mechanisms through 3D NMR. Firstly, a commonly used 3D NMR pulse sequence, tri-window pulse sequence, in actual oilfield was presented and its data processing workflow was exhibited briefly. Then, random-walk method was modified to simulate the pulse sequence measurements at different pore-fluid cases in tight sandstone, such as pure water case, oil-water case, and gas-water case. Finally, pore-fluid 3D NMR responses were exhibited and their microscopic mechanisms were explored. The results show that 3D NMR can identify different fluid components saturated in sedimentary rocks; wetting fluid saturation directly affect wetting fluid relaxation time, but the non-wetting fluid relaxation information is independent of fluid saturation; wetting fluid has an obvious restricted diffusion as a lower saturation; an obvious difference between gas signal and water or oil signal in two-dimensional relaxation spectra due to high longitudinal to transverse relaxation time ratio of pore gas. These simulations provide a theoretical basis for interpreting 3D NMR macroscopic responses, which should be helpful for pore-fluid identifying in tight oil and gas reservoirs.

A novel pulse sequence and inversion algorithm of three-dimensional low field NMR technique in unconventional resources.

Compared with two-dimensional (2D) nuclear magnetic resonance (NMR) technique like correlations among the transversal relaxation time (T2), the longitudinal relaxation time (T1), and the diffusion coefficient correlation (D), three-dimensional (3D) NMR technique is superior with the complete measurement of T2, T1, and D simultaneously. It can solve the problem of overlaps in 2D correlation map and is helpful to characterize relaxation components in unconventional resources such as tight gas and oil shale. However, the existed 3D NMR technique is restricted due to the loss of short relaxation information and the inversion inaccuracy that caused by the incomplete measurement of the diffusion editing window. We developed a tri-window pulse sequence to collect the full decaying information of porous media. In the first window, the inversion-recovery pulse sequence is applied for T1 encoding. In the second window, D and T2 are encoded by an adjustable continuous pulse field gradient and echo spacing (TE). In the last window, CPMG with the shortest TE is used to acquire diffusion-free relaxation information. We then proposed a joint inversion algorithm named "composite-data-processing" to obtain the 3D correlation map. The algorithm adopts the dimension reduction technique and the truncated singular value decomposition (TSVD) to speed up the inversion process and enhance the inversion stability. Numerical simulations show that good estimations of the inversion results are obtained at different signal to noise ratios (SNRs). Our results suggest that the novel pulse sequence and inversion algorithm of 3D NMR can be effectively applied to the exploration of unconventional resources.

Helical Structure of Recombinant Melittin.

Melittin is an extensively studied, 26-residue toxic peptide from honey bee venom. Because of its versatility in adopting a variety of secondary (helix or coil) and quaternary (monomer or tetramer) structures in various environments, melittin has been the focus of numerous investigations as a model peptide in protein folding studies as well as in studies involving binding to proteins, lipids, and polysaccharides. A significant body of evidence supports the view that melittin binds to these macromolecules in a predominantly helical conformation, but detailed structural knowledge of this conformation is lacking. In this report, we present nuclear magnetic resonance (NMR)-based structural insights into the helix formation of recombinant melittin in the presence of trifluoroethanol (TFE): a known secondary structure inducer in peptides. These studies were performed at neutral pH, with micromolar amounts of the peptide. Using nuclear Overhauser effect (NOE)-derived distance restraints from three-dimensional NMR spectra, we determined the atomic resolution solution NMR structure of recombinant melittin bearing a TFE-stabilized helix. To circumvent the complications with structure determination of small peptides with high conformational flexibility, we developed a workflow for enhancing proton NOEs by increasing the viscosity of the medium. In the TFE-containing medium, recombinant monomeric melittin forms a long, continuous helical structure, which consists of the N- and C-terminal α-helices and the noncanonical 310-helix in the middle. The noncanonical 310-helix is missing in the previously solved X-ray structure of tetrameric melittin and the NMR structure of melittin in methanol. Melittin's structure in TFE-containing medium provides insights into melittin's conformational transitions, which are relevant to the peptide's interactions with its biological targets.

Surgical treatment of trigeminal neuralgia with no neurovascular compression: A retrospective study and literature review

The objective of the current study is to summarize the experiences of 360-degree circumferential arachnoid dissection for TN in patients without NVC. Clinical data from 19 TN patients without NVC who underwent 360-degree circumferential arachnoid dissection were retrospectively analyzed. All patients underwent preoperative examinations of three-dimensional time-of-flight magnetic resonance angiography (3D-TOF-MRA) and three-dimensional nuclear magnetic resonance of constructive interference in steady state (3D-CISS) to identify the neurovascular structures around the trigeminal nerve. Surgical outcomes were evaluated based on the Barrow Neurological Institute (BNI) Pain Intensity Scale, and severity of pain was determined using the visual analogue scale (VAS) score. Preoperative imaging showed that no patients had NVC compression. The preoperative BNI pain score was IV among 9 patients and V among other 10 patients. The preoperative VAS score was 7.89 ± 0.658. After the operation, pain disappeared in all 19 patients, and transient facial numbness occurred in 3 patients. During follow-up, pain in 2 of 19 patients (10.5%) recurred at postoperative 20 and 23 months. This study demonstrated that 360-degree circumferential arachnoid dissection is an alternative treatment for trigeminal neuralgia in patients with no neurovascular compression, especially when a thickened arachnoid, angulation, or torsion of the trigeminal nerve root was found during the operation. However, further study is needed to compare the outcomes of different surgical strategies and prove the efficacy of 360-degree circumferential arachnoid dissection in cases without neurovascular compression (NVC).

A Hybrid Method for NMR Data Compression Based on Window Averaging (WA) and Principal Component Analysis (PCA)

Prior to the advent of nuclear magnetic resonance (NMR) data inversion, a common approach for handling the large amount of raw echo data collected by NMR logging was data compression for improving the inversion speed. A fast compression method with a high compression ratio is required for processing NMR logging data. In this paper, we proposed a hybrid method to compress NMR data based on the window averaging (WA) and principal component analysis (PCA) methods. The proposed method was compared with the WA method and the PCA method in terms of the compression times of simulated one-, two-, and three-dimensional NMR data, the inversion times of compressed echo data, and the accuracy of NMR maps created with and without compression. We processed NMR log data and compared the inversion results with different compression methods. The results indicated that the proposed method with a high compression speed and a high compression ratio can be used for NMR data compression, and its accuracy depended on the precompressed echo number, and it is obvious that the method have practical applications for NMR data processing, especially for multi-dimensional NMR.

Identification and Rationalization of Kinetic Folding Intermediates for a Low-Density Lipoprotein Receptor Ligand-Binding Module.

Many mutations that cause familial hypercholesterolemia localize to ligand-binding domain 5 (LA5) of the low-density lipoprotein receptor, motivating investigation of the folding and misfolding of this small, disulfide-rich, calcium-binding domain. LA5 folding is known to involve non-native disulfide isomers, yet these folding intermediates have not been structurally characterized. To provide insight into these intermediates, we used nuclear magnetic resonance (NMR) to follow LA5 folding in real time. We demonstrate that misfolded or partially folded disulfide intermediates are indistinguishable from the unfolded state when focusing on the backbone NMR signals, which provide information on the formation of only the final, native state. However, 13C labeling of cysteine side chains differentiated transient intermediates from the unfolded and native states and reported on disulfide bond formation in real time. The cysteine pairings in a dominant intermediate were identified using 13C-edited three-dimensional NMR, and coarse-grained molecular dynamics simulations were used to investigate the preference of this disulfide set over other non-native arrangements. The transient population of LA5 species with particular non-native cysteine connectitivies during folding supports the conclusion that cysteine pairing is not random and that there is a bias toward certain structural ensembles during the folding process, even prior to the binding of calcium.

A novel efficient inversion method for three-dimensional NMR and the optimization of activation sequences and acquisition parameters

A three-dimensional (3D) nuclear magnetic resonance (NMR) spectrum can simultaneously provide distributions of longitudinal relaxation time (T1), transverse relaxation time (T2), and diffusivity (D); thus, it greatly improves the capacity of fluid identification, typing, and quantitative evaluations. However, several challenges that significantly hinder the widespread application of this technique remain. The primary challenges are the high time and memory costs associated with the current 3D NMR inversion algorithms. In addition, an activation sequence optimization method for 3D NMR inversions has not been developed. In this paper, a novel inversion method for 3D NMR spectra and a detailed optimization method for activation sequences and acquisition parameters were proposed. The novel method, namely randomized singular value decomposition (RSVD) inversion algorithm, can reduce memory requirements and ensure computational efficiency and accuracy. Window averaging (WA) technique was also adopted in this study to further increase computational speed. The optimized method for pulse sequences is mainly based on projections of the 3D NMR spectra in the two-dimensional (2D) and one-dimensional (1D) domains. These projections can identify missing NMR properties of different fluids. Because of the efficiency and stability of this novel algorithm and the optimized strategy, the proposed methods presented in this paper could further promote the widespread application of 3D NMR.

A heteronuclear intermolecular single-quantum coherence scheme for high-resolution 2D J-resolved 1H NMR spectra in inhomogeneous magnetic fields

ABSTRACTBased on heteronuclear intermolecular single-quantum coherences between proton (1H) and quadrupolar nuclei (i.e. deuterium 2H), a three-dimensional nuclear magnetic resonance (NMR) pulse sequence is proposed for recovering high-resolution two-dimensional J-resolved NMR spectra from samples mixed with a deuterated solvent in the presence of large magnetic field inhomogeneities. Benefitting from excitation of spins via two different radio frequency (RF) transmit channels, this sequence is suitable for applications in randomly large inhomogeneous fields and the solvent suppression generally required in homonuclear intermolecular multiple-quantum coherence approaches is no longer necessary. Systematic theoretical analyses are given based on the distant dipolar field treatment. Experiment on a sample of corn oil in deuterated acetone and ethyl 3-bromopropionate and acetone dissolved in DMSO-d6 in a deshimmed field with severe inhomogeneous broadening is performed to show the feasibility and applicability of this sequence.

Identification, Characterization, and Three-Dimensional Structure of the Novel Circular Bacteriocin, Enterocin NKR-5-3B, from Enterococcus faecium.

Enterocin NKR-5-3B, one of the multiple bacteriocins produced by Enterococcus faecium NKR-5-3, is a 64-amino acid novel circular bacteriocin that displays broad-spectrum antimicrobial activity. Here we report the identification, characterization, and three-dimensional nuclear magnetic resonance solution structure determination of enterocin NKR-5-3B. Enterocin NKR-5-3B is characterized by four helical segments that enclose a compact hydrophobic core, which together with its circular backbone impart high stability and structural integrity. We also report the corresponding structural gene, enkB, that encodes an 87-amino acid precursor peptide that undergoes a yet to be described enzymatic processing that involves adjacent cleavage and ligation of Leu(24) and Trp(87) to yield the mature (circular) enterocin NKR-5-3B.

Discrete decoding based ultrafast multidimensional nuclear magnetic resonance spectroscopy.

The three-dimensional (3D) nuclear magnetic resonance (NMR) spectroscopy constitutes an important and powerful tool in analyzing chemical and biological systems. However, the abundant 3D information arrives at the expense of long acquisition times lasting hours or even days. Therefore, there has been a continuous interest in developing techniques to accelerate recordings of 3D NMR spectra, among which the ultrafast spatiotemporal encoding technique supplies impressive acquisition speed by compressing a multidimensional spectrum in a single scan. However, it tends to suffer from tradeoffs among spectral widths in different dimensions, which deteriorates in cases of NMR spectroscopy with more dimensions. In this study, the discrete decoding is proposed to liberate the ultrafast technique from tradeoffs among spectral widths in different dimensions by focusing decoding on signal-bearing sites. For verifying its feasibility and effectiveness, we utilized the method to generate two different types of 3D spectra. The proposed method is also applicable to cases with more than three dimensions, which, based on the experimental results, may widen applications of the ultrafast technique.

Multidimensional High-Resolution Magic Angle Spinning and Solution-State NMR Characterization of (13)C-labeled Plant Metabolites and Lignocellulose.

Lignocellulose, which includes mainly cellulose, hemicellulose, and lignin, is a potential resource for the production of chemicals and for other applications. For effective production of materials derived from biomass, it is important to characterize the metabolites and polymeric components of the biomass. Nuclear magnetic resonance (NMR) spectroscopy has been used to identify biomass components; however, the NMR spectra of metabolites and lignocellulose components are ambiguously assigned in many cases due to overlapping chemical shift peaks. Using our 13C-labeling technique in higher plants such as poplar samples, we demonstrated that overlapping peaks could be resolved by three-dimensional NMR experiments to more accurately assign chemical shifts compared with two-dimensional NMR measurements. Metabolites of the 13C-poplar were measured by high-resolution magic angle spinning NMR spectroscopy, which allows sample analysis without solvent extraction, while lignocellulose components of the 13C-poplar dissolved in dimethylsulfoxide/pyridine solvent were analyzed by solution-state NMR techniques. Using these methods, we were able to unambiguously assign chemical shifts of small and macromolecular components in 13C-poplar samples. Furthermore, using samples of less than 5 mg, we could differentiate between two kinds of genes that were overexpressed in poplar samples, which produced clearly modified plant cell wall components.

Structure-based functional identification of Helicobacter pylori HP0268 as a nuclease with both DNA nicking and RNase activities.

HP0268 is a conserved, uncharacterized protein from Helicobacter pylori. Here, we determined the solution structure of HP0268 using three-dimensional nuclear magnetic resonance (NMR) spectroscopy, revealing that this protein is structurally most similar to a small MutS-related (SMR) domain that exhibits nicking endonuclease activity. We also demonstrated for the first time that HP0268 is a nicking endonuclease and a purine-specific ribonuclease through gel electrophoresis and fluorescence spectroscopy. The nuclease activities for DNA and RNA were maximally increased by Mn2+ and Mg2+ ions, respectively, and decreased by Cu2+ ions. Using NMR chemical shift perturbations, the metal and nucleotide binding sites of HP0268 were determined to be spatially divided but close to each other. The lysine residues (Lys7, Lys11 and Lys43) are clustered and form the nucleotide binding site. Moreover, site-directed mutagenesis was used to define the catalytic active site of HP0268, revealing that this site contains two acidic residues, Asp50 and Glu54, in the metal binding site. The nucleotide binding and active sites are not conserved in the structural homologues of HP0268. This study will contribute to improving our understanding of the structure and functionality of a wide spectrum of nucleases.