Preprints  

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

Providing evidence that proteins are dynamical molecules is a major contribution of solution NMR to structural biology. But the classical experiments to measure dynamics can only be applied to small proteins. This is regretful, because larger proteins often display conformational changes in which dynamics plays a functional role. We show here that dynamic information can be extracted from HSQC NMR spectra using a computational approach. Such spectra are widely available also for larger proteins.

Unlike rigid folded proteins, intrinsically disordered proteins (IDPs) exist as an ensemble of dynamically interconverting structures. Cross-correlated spin-relaxation experiments are particularly suited to characterize the structural dynamics of IDPs as they encode a protein's "structural memory". By translating diffusion models of folded proteins to segmental motions in IDPs, we define an unambiguous experimental measure for the presence of anisotropic dynamics in the protein backbone.

We develop an exact analytical description of spinning sideband intensities in magic-angle spinning NMR, e.g. for ¹³C CSA. This can be used in spectral fitting to obtain tensor parameters, and plays out its advantage in the analysis of orientation effects in non-isotropic samples. We present an improved approach to process data obtained with the simple “syncMAS” experiment, which is powerful but not very popular due to the so far not very transparent and difficult data analysis.

Kidney stones (KS) are a major health problem in industrialized countries. The study of KS is presently at the heart of a concerted multi-disciplinary axis of research involving physicians, physical chemists and spectroscopists. In this contribution, an in-depth structural description of kidney stones is proposed by implementing a combination of multinuclear and multidimensional solid state NMR methodology.

Long-lived nuclear spin order overcomes the limited timescale of polarization storage by displaying much longer lifetimes compared to the longitudinal and transversal magnetizations. However, the excitation of these symmetry-adapted states required spin choreographies that were often overshadowed by their applications. Here, we recapitulate some of the milestones achieved in designing suited pulse sequences and comment on some future perspectives.

NMR spectroscopy is at the front row of the analytical methods used in organic chemistry and phytochemistry. Multiplet structure deconvolution has been revisited in the perspective of a robust integration in the computer-assisted workflow of 1D spectra analysis. New features include the management of coupling partners with spin > 1/2, second-order effects and partial signal overlap.

The RFDR sequence has been widely used for homonuclear recoupling. The article 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 sequences. Such analysis allows better understanding of the influence of offsets, paths of magnetization transfers for both these experiments, as well as the crucial role of XY phase cycling.

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 -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).

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.

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 to an iridium catalyst. In this work we present an approach to measure NMR relaxation as well as rate constants of para-hydrogen association/dissociation to the iridium catalyst. These rates are important to understand the different PHIP efficiency observed for different substrate molecules.

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.

A straightforward lineshape analysis of the ¹³C spectral asymmetries of [2-¹³C]sodium acetate can be used to indirectly evaluate the ¹H polarization of the CH₃ group, and likely entire samples in the case of rapid and homogeneous ¹H-¹H spin diffusion. The results are potentially advantageous for polarizers that lack ¹H radiofrequency hardware, measurements which are influenced by radiation damping or large background signals, or where acquisition of thermal equilibrium spectra are not feasible.

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.

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 NMR signals which may be used, amongst other things, for imaging of metabolism in the human body.

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.

NMR studies can be of great help to understand the molecular mechanisms of nucleosome function. 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.

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 lysinyl 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.

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.

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 that it may be affected by the electrochemical equipment itself.

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, both with 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.

The overall aim of the study is to provide a basis from which to improve the ability of tamoxifen family drugs to reduce the activity of a secondary target protein, calmodulin, during tumour development. The main conclusion is that the binding of a tamoxifen analogue is quite unlike that of other anti-calmodulin compounds, in that two drug molecules bring the two domains of calmodulin into close proximity, but they are not fixed in orientation relative to the protein.

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.

We present a system for facilitated sample vitrification, melting, and transfer in dissolution dynamic nuclear polarization experiments. The system enables inserting 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.

DEER is a method, which provide information on the structure of proteins by attaching to the protein two spins labels at 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. In this work we show how to set up the experiment for best sensitivity when the solvent is water, H₂O.

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 an regulatory mechanism.

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.

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 others [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.

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.

Shaped radiofrequency pulses are widely used in NMR spectroscopy to provide selective excitation or inversion of magnetization. Efficient accurate methods for calculating the performance of such pulses enable understanding of existing pulses and facilitate optimization of 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 also will be useful in other areas of theoretical NMR spectroscopy.

We show here that information about protein molecular dynamics can be directly extracted out of a single NMR spectrum. Understanding protein dynamics is essential to understanding its function, energetics and biology. We hope that this contribution helps bring this insight to a much broader application field and audience community than previously possible.

The demonstration of the Finite Pulse Radio Frequency Driven Recoupling experiment was firstly published over the past 20 years. Since it has become one of the used experiments. In this article we consider that experiment using other theoretical and simulated approaches with respect to the previous works. These approaches help us to understand more clearly the influence of each part of that experiment and to see how exactly the spin system is evaluated during it.

We recently added 2D NMR spectroscopy to the well-established techniques able to obtain information on the process of unfolding using resonances of residues in the hydrophobic core of a protein. We asked whether a detailed analysis of the individual stability curves could provide additional site-specific information. By monitoring both cold and heat denaturation of individual residues we convincingly demonstrate that the two unfolding processes are intrinsically different.

Nowadays, almost all MRI technique used in clinical applications are based on a Fourier transformation of the raw data. Alternative non-Fourier magnetic resonance imaging techniques can offer some advantages under certain conditions. SPatio-temporal ENcoding (SPEN) is a non-Fourier method that usually employs frequency-swept radio frequency pulses and can give good images in inhomogeneous magnetic fields. In this work we show how one can improve this technique by using quadratic gradients.