A new way is presented for creating lanthanide binding sites on proteins using site-specifically introduced phosphoserine residues. The paramagnetic effects of lanthanides generate long-range effects, which contain structural information and are readily measured by NMR spectroscopy. Excellent correlations between experimentally observed and back-calculated pseudocontact shifts attest to very good immobilization of the lanthanide ions relative to the proteins.

A method is demonstrated that can help in sequence-specific NMR signal assignment to nuclear spins near a strongly paramagnetic metal in an enzyme. A combination of paramagnetically tailored NMR experiments and second-shell mutagenesis was used to attribute previously observed chemical exchange processes in the active site of laccase to specific histidine ligands. The signals of nuclei close to the metal can be used as spies to unravel the role of motions in the catalytic process.

The efficiency of MRI contrast agents can increase by exploiting magnetization transfer effects occurring in slow-rotating nanoparticles containing a paramagnetic metal ion and a large number of exchangeable surface protons, which can increase the water proton relaxation rate. Occurrence of magnetization transfer should also be considered to determine accurate metal–proton distances from the experimental proton relaxation rates for protons farther than 15 Å from the paramagnetic metal.

We show ¹³C spin hyperpolarization using laser irradiation at room temperature in two different systems: 1. pentacene doped in organic crystals of benzoic acid and 2. nitrogen-vacancy (NV) color centers in microdiamonds. The ¹³C NMR signal was enhanced by a factor of > 3000 using pentacene and > 300 using NV centers as polarization sources. We analyze the pros and cons of these two systems and discuss their prospects for room-temperature enhancement of NMR signals.

Modern contrast agents for MRI lead to new methods for e.g. cancer diagnosis. One of these promising methods is called PHIP (parahydrogen-induced polarization) and can be performed at low instrumentation costs. However, it is based on parahydrogen which has to be produced on-site. Since commercial generators cost up to 150 000 Euro, we decided to provide the blueprints of our generator (< 3000 Euro) as open source to enable more groups to build their own generator and enter this research field.

While most proteins adopt one conformation, several interconvert between two or more very different structures. Knowing how sequence changes and small-molecule binding can control this behavior is essential for both understanding biology and inspiring new “molecular switches” which can control cellular pathways. This work contributes by examining these topics in the ARNT protein, showing that features of both the folded and unfolded states contribute to the interconversion process.

In this work we explore how magnetic resonance information can be obtained from simple field dependencies of reaction yields for recombining radical pairs, without actually driving any resonance transitions. An ESR spectrum can be observed in the region of relatively weak but nonzero applied field due to interference in the coherently created spin system of the radical pair. We also discuss connections between the level (anti)crossing and conventional magnetic resonance spectroscopy.

Catalytic hydrogenation of unsaturated molecular precursors with parahydrogen provides a major signal enhancement in NMR, which is vital for many technical as well as biomedical applications. We show that dilution of catalytically active Pd with a less active metal in heterogeneous Pd-based catalysts provides significantly larger enhancements and that the judicious choice of the dilution metal can further improve the performance, as revealed by comparing Pd-Ag and Pd-In bimetallic catalysts.

The application of anisotropic NMR parameters for the correct structural assignment of small molecules requires the use of partially ordered media. Previously, we demonstrated that P3D simulations allow for the determination of the correct diastereomer. Here we show that when steric or electrostatic factors dominate the alignment, P3D retains its diastereomer discrimination power and can define conformational ensembles of flexible molecules from sparse NMR data.

Typically, low-field Overhauser dynamic nuclear polarization (ODNP) experiments are 1D NMR experiments to study hydration dynamics. Here, we demonstrate the application of ODNP-enhanced 2D J-resolved (JRES) spectroscopy to improve spectral resolution beyond the limit imposed by the line broadening introduced by the paramagnetic polarizing agent. Crucial to these experiments is interleaved spectral referencing to compensate for temperature-induced field drifts over the course of the experiment.

NMR study of large proteins such as SARS-CoV-2 Main Protease can be challenging when exchange broadening, multiple stable conformations, and back-exchanging the fully deuterated chain pose problems. We demonstrate that 4D NMR, including an extension of 3D NOE-NOE spectroscopy, provides an effective tool for spectral analysis. In combination with X-ray coordinates, the 4D NMR data are particularly useful for extending and validating assignments and for probing structural features.

We report data on nuclear polarization formed in photoreaction of intramolecular electron transfer from adenine to flavin that was revealed several years ago by R. Kaptein by CIDNP. The question of whether one or two types of biradicals are formed stays unresolved. This work clarified the problem and confirmed that single common emissive maximum in magnetic field dependence of CIDNP may originate from single level crossing of short-lived biradical and that the FAD molecule has a compact shape.

The NMR-MOUSE is a magnetic resonance tool for non-destructive materials testing inside a laboratory. The history and use of this sensor are reviewed with attention to issues encountered when employed outside. Improvements are outlined to facilitate outdoor measurements.

Double electron–electron resonance (DEER) provides information on the structure of proteins by attaching two spin labels to the protein at a well-defined location and measuring the distance between them. The sensitivity of the method in terms of the amount of the protein that is needed for the experiment depends strongly on the relaxation properties of the spin label and the composition of the solvent. We show how to set up the experiment for best sensitivity when the solvent is water (H₂O).

Shaped radiofrequency pulses are widely used in nuclear magnetic resonance (NMR) spectroscopy for selective excitation or inversion of magnetization. Efficient, accurate methods for calculating the performance of pulses enable the understanding of existing pulses and help to optimize new pulses. A new approach for approximating the effects of shaped pulses is introduced and applied to some popular shaped pulses as examples. This approach will also be useful in other areas of NMR spectroscopy.

NMR studies can be of great help in understanding the molecular mechanisms of nucleosome functions. For solid-state NMR, nucleosomes need to be tightly packed together. We show that centrifugation of nucleosomes results in formation of gels with very high packing ratios yet without pronounced order in the packing and without formation of specific or stable inter-nucleosome contacts. This makes the approach suitable also for the study of proteins that bind weakly to the nucleosome.

The iron–sulfur cluster binding properties of human mitoNEET have been investigated by 1D and 2D 1H paramagnetic NMR spectroscopy. The NMR spectra of both oxidized and reduced mitoNEET are significantly different from those reported previously for other [Fe2S2] proteins. Our findings revealed the unique electronic properties of mitoNEET and suggests that the specific electronic structure of the cluster possibly drives the functional properties of different [Fe2S2] proteins.

Although both the hydrophobic aliphatic chain and hydrophilic ζ-amino group of the lysine side chain presumably contribute to the structures and functions of proteins, the dual nature of the lysine residue has not been fully understood yet, due to the lack of appropriate methods to acquire comprehensive information on its long consecutive methylene chain at the atomic scale. We describe herein a novel strategy to address the current situation using nuclear magnetic resonance spectroscopy.

Dynamic nuclear polarization (DNP) magic angle spinning NMR yields huge signal enhancements and is attractive for signal-limited cases. We studied polarization transfer pathways underlying sensitivity and resolution in DNP experiments on tubular assemblies of HIV-1 CA protein. All three polarization transfer pathways are simultaneously active and can be selected by carefully choosing experimental conditions. Our study highlights the unique capabilities of DNP for structural biology applications.

NMR relaxation rate constants are powerful probes of the functional dynamics of biological macromolecules, such as proteins and nucleic acids. However, variations in choice of models used to analyze relaxation data obscure variations due to critical biological or chemical effects. Bootstrap aggregation, a recently developed statistical method, circumvents the model selection problem and enables more consistent and insightful interpretation of NMR relaxation data.

This work presents a setup for the investigation of electrochemical processes during operation (in operando) using nuclear magnetic resonance (NMR) spectroscopy. The setup was designed to minimize the interferences between the NMR instrument and the electrochemical equipment. Employing this setup, the dynamic equilibrium of carbon dioxide in aqueous bicarbonate electrolyte has been monitored in operando, revealing intercations with the electrode setup.

Methyl groups are rarely acidic. But in a lumazine derivative with biological significance the position-7 methyl group can be deprotonated. The existence of two protonation states is a prerequisite for a non-classical disproportionation reaction initiated by light. We provide evidence for such dismutation using a variant of nuclear magnetic resonance that is capable of detecting nuclear hyperpolarization. Our findings may shed new light on enzymatic riboflavin biosynthesis.

The folding, refolding and misfolding of biomacromolecules including proteins, DNA and RNA is an important area of biophysical research to understand functional and disease states of a cell. NMR spectroscopy provides detailed insight, with both high time and atomic resolution. These experiments put stringent requirements on signal-to-noise for often irreversible folding reactions. The review describes methodological approaches and highlights key applications.

Derivatives of the amino acids tryptophan and tyrosine were studied in their potential to enhance their signal in nuclear magnetic resonance spectroscopy (NMR) by laser-induced dynamic nuclear polarisation (CIDNP). Signal enhancement up to two orders of magnitude was obtained.

Reversible para-hydrogen-induced polarization (PHIP) is a technique to enhance NMR signals in solution, based on the association/dissociation of para-hydrogen and substrate molecules with/from an iridium catalyst. In this work we present an approach to measure NMR relaxation as well as rate constants of para-hydrogen association/dissociation with/from the iridium catalyst. These rates are important for understanding the different PHIP efficiencies observed for different substrate molecules.

This is an obituary for Konstantin Lvovitch Ivanov.

The RFDR sequence has been widely used for homonuclear recoupling. The paper describes a heteronuclear version of RFDR. HET-RFDR sequence transfers longitudinal polarization between heteronuclear pairs by applying RFDR on two channels simultaneously. We perform an operator analysis of HET-RFDR and RFDR. Such an analysis allows for better understanding of the influence of offsets and paths of magnetization transfers for both these experiments, as well as the crucial role of XY phase cycling.

The Wnt signalling pathway plays a major role in prevention of cancer, whereby the protein Dishevelled connects from the transmembrane receptor Frizzled to downstream effectors via its PDZ domain. Here, cycles of chemical synthesis and structural biology are applied to develop PDZ ligands that block the Frizzled–Dishevelled interaction using NMR for screening, in ligand development, and for deriving structure–activity relationships. Cellular reporter assays demonstrate their efficacy.

Trigger factor (TF) is a conserved bacterial molecular chaperone that exists in a monomer–dimer equilibrium in solution. It binds to the ribosome as a monomer to facilitate folding of nascent polypeptide chains. We showed that dimeric TF exhibits distinct domain dynamics and conformational polymorphism and that TF contains multiple substrate binding sites that are only accessible in its monomeric form. The equilibrium of TF in different oligomeric states may serve as a regulatory mechanism.

We present a system for facilitated sample vitrification, melting, and transfer in dissolution dynamic nuclear polarization experiments. The system enables insertion of a sample through an airlock in combination with a dedicated dissolution system that is inserted through the same airlock shortly before the melting event. The cryostat can thus be operated continuously at low temperatures, and the melting process is rapid as no pressurization steps of the cryostat are necessary for dissolution.

Magnetic resonance experiments are described using a mathematical object called the spin density operator. The article explores the regions of an abstract space called Liouville space which may be occupied by the density operator. The study is of particular relevance to hyperpolarized magnetic resonance, which is capable of generating greatly enhanced nuclear magnetic resonance signals which may be used, amongst other things, for imaging of metabolism in the human body.

The use of hyperpolarized species, i.e., species whose nuclear polarization is transiently multiplied by several orders of magnitude, can perfectly be combined with the use of benchtop NMR spectrometers. In particular, laser-polarized ¹²⁹Xe, due to its large chemical shift range and the proximity of its resonance frequency to ¹³C, can be observed on a non-dedicated low-field spectrometer. Detection of low amounts of molecules can be achieved using NMR techniques based on chemical exchange.

An approach is described for formulating the kinetic master equations of the time evolution of hyperpolarized NMR signals in reacting (bio)chemical systems. We take a stepwise approach to formulate mathematical models of enzyme systems that agree with standard descriptions of (bio)chemical kinetics while remaining capable of describing the time evolution of magnetization described by the Bloch–McConnell equations.

The transfer of polarization from protons to half-integer spin quadrupolar nuclei provides insights into the proximities between these nuclei. We analyse the best transfers at 20 and 62 kHz MAS obtained with RINEPT and PRESTO recouplings using adiabatic or composite π pulses. The 1H magnetization is best transferred using PRESTO-R22,7,2 (180) or -R16,6,7 (270,90) at 20 or 62 kHz MAS, respectively, and RINEPT-SR4,2,1 (270,90).

This publication presents research in biomolecular NMR, spin hyperpolarisation and spin chemistry. Robert Kaptein made key contributions to magnetic resonance and proposed the radical pair mechanism for chemically induced dynamic nuclear polarisation (CIDNP). He developed laser CIDNP and computational methods for protein structure determination. He is known for studying the lac repressor and its DNA complexes. He played a leading role in establishing the NMR large-scale facilities in Europe.

Isotope filters are pivotal for studying interactions between labelled and unlabeled moieties by NMR. We present an isotope filter method to complement existing filters and attenuate undesired signals that escaped them. This advance will readily facilitate molecular studies of binding events or post-translational modifications by NMR.

In drug discovery, ready access to accurate atomic resolution information revealing conformations and dynamics of novel small molecules in solution guides rational design to optimize drug potency and physical chemistry. We developed an efficient integration of molecular dynamics calculations and visualization protocols with NMR conformational analysis of free ligands in solution. Agreement between experiment and theory inspires confidence in prospective computational analysis of new designs.

Sample rotation around the magic angle averages out the dipolar couplings in homonuclear spin systems in a first-order approximation. However, in higher orders, residual coupling terms remain and lead to a broadening of the spectral lines. We investigate the source of this broadening and the effects on the powder line shape in small spin systems with and without chemical shifts. We show that one can expect different scaling behavior as a function of the spinning frequency for the two cases.

NMR represents a key spectroscopic technique for studying intrinsically disordered proteins (IDPs) that lack a stable three-dimensional structure. We present a set of NMR experiments tailored for proline residues, which are highly abundant in IDPs. The novel experiments have very interesting properties for the investigation of IDPs of increasing complexity.

The radio-frequency (rf) field amplitude in solid-state NMR probes changes over the sample volume, i.e. different parts of the sample will experience different nutation frequencies. If the sample is rotated inside the coil as it is typical for magic angle spinning in solid-state NMR, such a position-dependent inhomogeneity leads to an additional time dependence of the rf field amplitude. We show that such time-dependent modulations do not play an important role in many experiments.