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International audience; Doping of transition metals within a semiconductor quantum dot (QD) has a high impact on the optical and magnetic properties of the QD. In this study, we report the synthesis of Mn 2+-doped Ag−In−Ga−Zn−S (Mn:AIGZS) QDs via thermolysis of a dithiocarbamate complex of Ag + , In 3+ , Ga 3+ , and Zn 2+ and of Mn(stearate) 2 in oleylamine. The influence of the Mn 2+ loading on the photoluminescence (PL) and magnetic properties of the dots are investigated. Mn:AIGZS QDs exhibit a diameter of ca. 2 nm, a high PL quantum yield (up to 41.3% for a 2.5% doping in Mn 2+), and robust photo-and colloidal stabilities. The optical properties of Mn:AIGZS QDs are preserved upon transfer into water using the glutathione tetramethylammonium ligand. At the same time, Mn:AIGZS QDs exhibit high relaxivity (r 1 = 0.15 mM −1 s −1 and r 2 = 0.57 mM −1 s −1 at 298 K and 2.34 T), which shows their potential applicability for bimodal PL/magnetic resonance imaging (MRI) probes.

International audience; Peptide-based hydrogels are physical gels formed through specific supramolecular self-assembling processes, leading to ordered nanostructures which constitutes the water entrapping scaffold of the soft material. Thanks to the inherent properties of peptides, these hydrogels are highly considered in the biomedical domain and open new horizons in terms of application in advanced therapies and biotechnologies. The use of one, and only one, native peptide to formulate a gel is by far the most reported approach to design such materials, but suffers from several limitations, including in terms of mechanical properties. To improve peptide-based hydrogels interest and give rise to innovative properties, several strategies have been proposed in the recent years, and the development of multicomponent peptide-based hydrogels appears as a promising and relevant strategy. Indeed, mixing two or more compounds to develop new materials is a much-used approach that has proven its effectiveness in a wide variety of domains, including polymers, composites and alloys. While still limited to a handful of examples, we would like to report herein on the formulation and the comprehensive study of multicomponent hybrid DNA-nucleobase/peptide-based hydrogels using a multiscale approach based on a large panel of analytical techniques (i.e., rheometry, proton relaxometry, SAXS, electronic microscopy, infrared, circular dichroism, fluorescence, Thioflavin T assays). Among the six multicomponent systems studied, the results highlight the synergistic role of the presence of the two complementary DNA-nucleobases (i.e., adenine/thymine and guanine/cytosine) on the coassembling process from structural (e.g., morphology of the nanoobjects) to physicochemical (e.g., kinetics of formation, fluorescence properties) and mechanical properties (e.g., stiffness, resistance to external stress). All the data confirm the relevance of the multicomponent peptide-based approach in the design of innovative hydrogels and bring another brick in the wall of the understanding of these complex and promising systems.

The emergence of a new generation of nanostructured materials opened a wide range of new potential applications for Bioglasses such as DNA vaccination, cellular treatment or drug delivery with a well-controlled loading and release of (bio)active molecules. In this study we compare structural, dynamic and thermal properties of water confined in two CaO–SiO2–P2O5 mesoporous bioactive glasses (MBGs). One of the MBGs is prepared by standard sol-gel methods and the other by microfluidic procedures, therefore both MBGs have the same molar compositions (92% SiO2, 6% CaO, and 2% P2O5) but different textural properties (surface areas, porous volume and grain architecture). These materials are intended to interact with body fluids which are essentially composed of water, it is therefore crucial to understand the water-MBG interaction. With this objective, we apply a complementary approach based on 1H field-cycling NMR relaxometry, 1H magic-angle spinning NMR spectroscopy and differential scanning calorimetry (DSC). With low-field relaxometry we observe that adsorbed water has the same behavior in the mesoporous channels of both systems, but that the escaping of water molecules towards the bulk is slightly faster in the microfluidic material. By combining 1H MAS NMR and DSC to study a series of MBG samples at variable temperature and water content, we observe that for low temperature and low water content, pooling of water molecules is less pronounced in the case of the microfluidic MBG. All our results therefore point towards a microcapsule organization of the microfluidic MBG that provides better water access to the full porous volume.

Attempts to prepare a supramolecular M4L2 box involving two tetra-bipyridyl calix[4]arene units and four copper(I) cations afforded, in fact, complex 2 for which, in the solid state, X-ray diffraction analysis showed that the stoichiometry is indeed M4L2, with the two calixarene subunits associated in a “hand-to-hand” mode by complexation via two adjacent bipyridine units to two copper(I) cations, and each group of two residual, adjacent, bipyridine arms is complexed to one copper(I) centre. Mass spectrometry, UV spectroscopy and preliminary 1H-NMR analysis are in preliminary accordance with a monocalixarenic right/left handed M2L structure. Nevertheless, some particularities of the NMR spectrum related to Ar-CH2-Ar and O-CH2bpy resonance signals, the possible generation of the M2L species during the mass spectrometry experiment and the lack of sensitivity of the UV-visible titration caused us to proceed to deeper investigations, looking for other associations, notably the M4L2 “hand-to-hand” systems. This involved 1D and 2D-NMR spectroscopies, among them 1H–15N HMBC, for the reconstruction of the molecule, but also the measurement of self-diffusion coefficients by NMR and electrochemistry. The 2D-NMR analysis and NMR self-diffusion evaluation were in accordance with the M2L structure, but the electrochemical self-diffusion experiment indicated a M4L2 structure.

Sodium nitroprusside (SNP), an active pharmaceutical ingredient, is encapsulated in biocompatible amorphous silica matrices and characterized by solid-state NMR. Through complementary NMR and pair distribution function (PDF) study, we previously showed that when confined in 1–2 nm sized nanopores of an amorphous silica matrix, the complex displays a “liquid like” isotropic motion (NMR) but still retains an associated anion/cation preferential arrangement (PDF). In this study, we present new insights into the physical state of the confined drug to explain the coexistence of these two seemingly contradictory characteristics. By performing temperature-dependent T1 relaxation experiments on the samples with different hydration levels, we retrieve respective motional activation energies for all three guest species inside the pores (sodium cations, nitroprusside anions, and water). For the hydrated samples, we identify a temperature range where results ascertain isotropic but correlated ionic motion of the SNP ...

International audience

At ambient temperature, conversion from 100% enriched para-hydrogen (p-H2; singlet state) to ortho-hydrogen (o-H2; triplet state) leads necessarily to the thermodynamic equilibrium proportions: 75% of o-H2 and 25% of p-H2. When p-H2 is dissolved in a diamagnetic organic solvent, conversion is very slow and can be considered as arising from nuclear spin relaxation phenomena. A first relaxation mechanism, specific to the singlet state and involving a combination of auto-correlation and cross correlation spectral densities, can be retained: randomly fluctuating magnetic fields due to inter-molecular dipolar interactions. We demonstrate here that (i) this dipolar mechanism is not sufficient for accounting for the para→ortho conversion rate, (ii) spin-rotation interaction, an intra-molecular mechanism, behaves similarly to random-field interaction and, thus, may be involved in the singlet relaxation rate. Also, as the para→ortho conversion is monitored by proton nuclear magnetic resonance (NMR) of dissolved o-H2 (p-H2 is NMR-silent), one has to account for H2 exchange between the liquid phase and the gas phase within the NMR tube, as well as for dissolution effects. Experimental evidence of the above statements is brought here in the case of two organic solvents: acetone-d6 and carbon disulfide. The observed temperature dependence of the para→ortho conversion rate shows that spin-rotation can be the dominant contribution to the p-H2 relaxation rate in the absence of tangible dipolar interactions. Our findings shed new light on the “mysterious” mechanism of the para→ortho conversion which has been searched for several decades.

Water in mesoporous materials possessing a two-dimensional hexagonal structure has been studied by the variation of its NMR longitudinal relaxation time T(1) as a function of the static magnetic field value, or equivalently of the NMR measurement frequency. This technique, dubbed relaxometry, has been applied from 5 kHz (measurement frequency) up to 400 MHz with various instruments including a variable-field spectrometer operating between 8 and 90 MHz. Moreover, the range 0-5 kHz could be investigated by transverse relaxation, T(2) denoting the corresponding relaxation time, and relaxation in the rotating frame, T(1ρ) denoting the corresponding relaxation time. Measurements of proton relaxation rates (inverse of relaxation times) have been performed with H(2)O and HOD (residual protons of heavy water) at water volumes of 80%, 60%, and 40% relative to the porous volume. Comparison between H(2)O and HOD shows clearly that, above 1 MHz where both sets of data are superposed, relaxation is purely intermolecular and due to paramagnetic relaxation (dipolar interactions of water protons with unpaired electrons of paramagnetic entities). Below 1 MHz, it is possible to subtract the intermolecular contribution (given by HOD data) from H(2)O data so that one is left with intramolecular relaxation which is solely due to water reorientational motions. The analysis of these low-frequency data (in terms of Lorentzian functions) reveals two types of water within the pores: one interacting strongly with the surface and the other corresponding to a second layer. High-frequency data, which arise from paramagnetic relaxation, exhibit again two types of water. Due to their correlation times, one type is assigned to relatively free water within the pores while the other type corresponds to bulk (interparticular) water. Their proportions, given as a function of the volume fraction, are consistent with the above assignments.

Heteronuclear Overhauser effect spectroscopy (HOESY) is a powerful method for tracking geometrical proximities between two heteronuclei (for instance, 1H and 13C, as this will be the case here). The method is based on cross-relaxation arising from dipolar interactions. Sensitivity permitting, it is applied in the 2D mode yielding all spatial correlations in a single experiment. Whenever sensitivity is not sufficient, it can be applied in the one-dimensional mode by selectively inverting one particular proton. In that case, it yields, from the carbon-13 spectrum, remote spatial correlations. The method has been employed here for the discrimination between two possible E or Z isomers in a medium-size molecule. Copyright © 2008 John Wiley & Sons, Ltd.

International audience; We report that at ambient temperature and with 100% enriched para-hydrogen (p-H-2) dissolved in organic solvents, paramagnetic spin catalysis of para -> ortho hydrogen conversion is accompanied at the onset by a negative ortho-hydrogen (o-H-2) proton NMR signal. This novel finding indicates an electron spin polarization transfer, and we show here that this can only occur if the H-2 molecule is dissociated upon its transient adsorption by the paramagnetic catalyst. Following desorption, o-H-2 is created until the thermodynamic equilibrium is reached. A simple theory confirms that in the presence of a static magnetic field, the hyperfine coupling between unpaired electrons and nuclear spins is responsible for the observed polarization transfer. Owing to the negative electron gyromagnetic ratio, this explains the experimental results and ascertains an as yet unexplored mechanism for para -> ortho conversion. Finally, we show that the recovery of o-H-2 magnetization toward equilibrium can be simply modeled, leading to the para -> ortho conversion rate.

International audience; A gradient-based sequence is proposed for efficiently filtering out all quantities except the longitudinal two-spin order, created by csa/dipolar cross-correlation rates. The dipolar interaction is between a proton and a heteronucleus, the csa being generally the one of the heteronucleus. Proton detection is carried out to benefit from maximum sensitivity. The resulting two-dimensional spectrum has the same aspect as an HMBC spectrum, implying the existence of a J coupling between the two considered nuclei, but here cross-peaks indicate the strength of the relevant csa/dipolar cross-correlation rate. The method is especially interesting in the case of medium-sized molecules where a given heteronucleus is subjected to several csa/dipolar cross-correlation effects, along with the corresponding J couplings.

We propose a method for the determination of (15)N csa/dipolar cross-correlation rates based on the measurement of the two apparent transverse (or longitudinal) relaxation rates associated with each component of the nitrogen doublet (N(alpha) and N(beta)). This is achieved by inserting a spin state selective scheme in conventional inverse Carr-Purcell-Meiboom-Gill (or inversion-recovery) pulse sequence which allows for the edition of a HSQC-type spectrum for each of the spin states. Transverse cross-correlation rates necessitate two independent sets of measurements (for N(alpha) and N(beta), respectively), whereas for longitudinal cross correlation rates, besides N(alpha) and N(beta) measurements, the method requires the knowledge of both the (15)N longitudinal auto-relaxation rate and the longitudinal two-spin order (2NzHz) auto-relaxation rate. These additional parameters are mandatory because of the non-exponential behavior of the N(alpha) and N(beta) longitudinal decays. Conversely, the present method does not require any complex manipulation of 2D spectra, the cross-correlation rates being obtained from the difference of the two (N(alpha) and N(beta)) apparent relaxation rates. This approach is applied to (15)N-labelled ubiquitin at two different magnetic fields (9.4 T and 14.1 T).

Comprehensive calculations are performed with the aim of arriving at a ready-to-use form for cross-correlation spectral density functions. Axial symmetry is assumed for tensors associated with the two interactions involved in the cross-correlation spectral density. Moreover, the derived formulas stem from the hypotheses of the “model-free” approach, i.e., a (generally fast) local motion superposed to a (generally slow) molecular tumbling. The restriction of fast local motions imposes the definition of two different order parameters S and S′ associated with the principal direction of the two considered interactions and it is the product of these order parameters, which is present in the general expression of a spectral density function. The latter includes two terms. The first one corresponds to fast local motions and involves the factor 1/2(3 cos2α − 1), where α is the angle made by the symmetry axes of the two considered tensors. The second term concerns only molecular tumbling sensed by a (virtual) director. Finally, it is shown that, consistently with the model-free approach, the product SS′ can be expressed as 1/2(3 cos2α − 1)S2 so that 1/2(3 cos2α − 1) factorizes the spectral density function. © 2003 Wiley Periodicals, Inc. Concepts Magn Reson Part A 19A: 65–70, 2003.

We investigate picosecond–nanosecond dynamics of the Rho-GTPase Binding Domain (RBD) of plexin-B1, which plays a key role in plexin-mediated cell signaling. Backbone 15N relaxation data of the dimeric RBD are analyzed with the model-free (MF) method, and with the slowly relaxing local structure/molecular dynamics (SRLS-MD) approach. Independent analysis of the MD trajectories, based on the MF paradigm, is also carried out. MF is a widely popular and simple method, SRLS is a general approach, and SRLS-MD is an integrated approach we developed recently. Corresponding parameters from the RBD dimer, a previously studied RBD monomer mutant, and the previously studied complex of the latter with the GTPase Rac1, are compared. The L2, L3, and L4 loops of the plexin-B1 RBD are involved in interactions with other plexin domains, GTPase binding, and RBD dimerization, respectively. Peptide groups in the loops of both the monomeric and dimeric RBD are found to experience weak and moderately asymmetric local ordering centered approximately at the C(i–1)(α)–C(i)(α) axes, and nanosecond backbone motion. Peptide groups in the α-helices and the β-strands of the dimer (the β-strands of the monomer) experience strong and highly asymmetric local ordering centered approximately at the C(i–1)(α)–C(i)(α) axes (N–H bonds). N–H fluctuations occur on the picosecond time scale. An allosteric pathway for GTPase binding, providing new insights into plexin function, is delineated.

Proton nuclear spin relaxation has been for the first time extensively used for a structural and dynamical study of low-molecular-weight organogels. The gelator in the present study is a modified phenylalanine amino acid bearing a naphthalimide moiety. From T(1) (spin-lattice relaxation time in the laboratory frame) and T(1ρ) (spin-lattice relaxation time in the rotating frame) measurements, it is shown that the visible gelator NMR spectrum below the liquid-gel transition temperature corresponds to a so-called isotropic compartment, where gelator molecules behave as in a liquid phase but exchange rapidly with the molecules constituting the gel structure. This feature allows one to derive, from accessible parameters, information about the gel itself. Nuclear Overhauser effect spectroscopy (NOESY) experiments have been exploited in view of determining not only cross-relaxation rates but also specific longitudinal rates. The whole set of relaxation parameters (at 25 °C) leads to a correlation time of 5 ns for gelator molecules within the gel structure and 150 ps for gelator molecules in the isotropic phase. This confirms, on one hand, the flexibility of the organogel fibers and, on the other hand, the likely presence of clusters in the isotropic phase. Concerning cross-relaxation rates, a thorough theoretical investigation in multispin systems of direct and relayed correlations in a NOESY spectrum allows one to make conclusions about contacts (around 2-3 A) not only between naphtalimide moieties of different gelator molecules but also between the phenyl ring and the naphtalimide moiety again of different gelator molecules. As a result, not only is the head-to-tail structure of amino acid columns confirmed but also the entangling of nearby columns by the naphthalimide moieties is demonstrated.

At very low or low frequency, the so-called dispersion curves (longitudinal relaxation rate as a function of the measurement frequency) are obtained with a ''relaxome- ter,'' which is generally run according to the ''field-cycling'' technique. Because of the very poor field homogeneity, relaxation times of a single (broad) signal are measured. When going to higher fields, with standard nuclear magnetic resonance spectrometers (possibly with variable field capacities), different lines are resolved. We show how the relaxation rates pertaining to these lines can be connected to the results obtained via the relaxometer. Among other things, we demonstrate why the magnitude mode (generally used for process- ing the relaxometer data) must be used with caution when several lines are resolved. Actually, we could obtain accurate 1 H dispersion curves in a very broad frequency range (5 kHz-400 MHz), and we show that a decomposition into several Lorentzian curves proves to be adequate, as far as paramagnetic relaxation due to dissolved oxygen is concerned. The method will be applied i) to nondegassed liquid toluene, ii) to the same solvent involved in the formation of an organogel. In that latter case, a low frequency contribution to the dispersion curve (possibly non-Lorentzian) reveals the part of the solvent within the organogel structure. The algorithm used for analyzing these dispersion curves is described. 2012 Wiley Periodicals, Inc. Concepts Magn Reson Part A 40A: 80-89, 2012.

Combinations of experimentally derived data from nuclear magnetic resonance spectroscopy and analyses of molecular dynamics trajectories increasingly allow us to obtain a detailed description of the molecular mechanisms by which proteins function in signal transduction. This chapter provides an introduction into these two methodologies, illustrated by example of a small GTPase–effector interaction. It is increasingly becoming clear that new insights are provided by the combination of experimental and computational methods. Understanding the structural and protein dynamical contributions to allostery will be useful for the engineering of new binding interfaces and protein functions, as well as for the design/in silico screening of chemical agents that can manipulate the function of small GTPase–protein interactions in diseases such as cancer.

An organogelation process depends on the gelator-solvent pair. This study deals with the solvent dynamics once the gelation process is completed. The first approach used is relaxometry, i.e., the measurement of toluene proton longitudinal relaxation time T(1) as a function of the proton NMR resonance frequency (here in the 5 kHz to 400 MHz range). Pure toluene exhibits an unexpected T(1) variation, which has been identified as paramagnetic relaxation resulting from an interaction of toluene with dissolved oxygen. In the gel phase, this contribution is retrieved with, in addition, a strong decay at low frequencies assigned to toluene molecules within the gel fibers. Comparison of dispersion curves of pure toluene and toluene in the gel phase leads to an estimate of the proportion of toluene embedded within the organogel (found around 40%). The second approach is based on carbon-13 T(1) and nuclear Overhauser effect measurements, the combination of these two parameters providing direct information about the reorientation of C-H bonds. It appears clearly that reorientation of toluene is the same in pure liquid and in the gel phase. The only noticeable changes in carbon-13 longitudinal relaxation times are due to the so-called chemical shift anisotropy (csa) mechanism and reflect slight modifications of the toluene electronic distribution in the gel phase. NMR diffusion measurements by the pulse gradient spin-echo (PGSE) method allow us to determine the diffusion coefficient of toluene inside the organogel. It is roughly two-thirds of the one in pure toluene, thus indicating that self-diffusion is the only dynamical parameter to be slightly affected when the solvent is inside the gel structure. The whole set of experimental observations leads to the conclusion that, once the gel is formed, the solvent becomes essentially passive, although an important fraction is located within the gel structure.

When a hydrogenation reaction is carried out with gaseous hydrogen enriched in its para- isomer in the earth magnetic field (prior to adiabatic insertion of the sample in the NMR magnet), enhanced proton longitudinal order (represented by 2 I z A I z B ) is created but also difference of enhanced polarizations ( I z A - I z B ). In a first part, it is shown theoretically and experimentally that the longitudinal relaxation time of this polarization difference is roughly twice the ones of individual polarizations. The second part is devoted to a pulse sequence designed for transforming this difference into net hyperpolarization. The evolution of this global hyperpolarization is studied experimentally in a third part and it is observed that a fraction of hyperpolarization possesses an effective longitudinal relaxation time similar to the one of the initial polarization difference. Those experimental results are interpreted by numerical calculations based on Solomon-type equations including the longitudinal order and possibly dipolar-csa cross correlation rates.

Owing to an extremely abundant literature making use of spin relaxation for structural studies, this review is limited to carbon-13 spectroscopy, to small or medium size molecules, to stereochemical and preferably geometrical determinations. The parameter of choice is evidently the Nuclear Overhauser effect (NOE) because it depends exclusively on the dipolar interaction mechanism, thus on 1/r6, where r is the distance between the two interacting spins. However, it depends also on the dynamical features of the system under investigation which must be characterized prior to any attempt for obtaining geometrical or stereochemical information. Therefore, this review is devoted not only to 1H–13C NOE but, more generally, to 13C longitudinal relaxation. After comprehensive theoretical developments, experimental methods presently available will be presented. The latter include the usual gated decoupling experiment and pulse experiments of the HOESY (Heteronuclear Overhauser Effect SpectroscopY) family. These pulse experiments, which imply carbon-13 observation, can be one-dimensional, selective one-dimensional or two-dimensional. The emphasis will be put on the interpretation which is different according to the occurrence or not of extreme narrowing conditions. Along with a literature survey, some selected examples will be presented in detail in order to illustrate the potentiality of the method.

Spin polarization transfer from parahydrogen (p-H(2)) to another molecular entity is generally thought to be mediated by longitudinal spin order (represented by the operator product I(z)(A)I(z)(B), A and B being the two hydrogen nuclei which originate from p-H(2) after a hydrogenation reaction). The longitudinal spin order leads to antiphase patterns in the proton NMR spectrum. In addition to these antiphase patterns, in-phase patterns, arising from polarization differences (represented by (I(z)(A)-I(z)(B))), have been experimentally observed. A complete theory, based on a density operator treatment, has been worked out and applied to the two types of parahydrogen induced polarization experiments: PASADENA (PArahydrogen and Synthesis Allow Dramatically Enhanced Nuclear Alignment; hydrogenation reaction inside the NMR magnet) and (ALTADENA) (Adiabatic Longitudinal Transport After Dissociation Engenders Nuclear Alignment; hydrogenation reaction outside the NMR magnet). It is shown that polarization differences are always created in the case of a PASADENA experiment but that their amplitude depends critically on the ratio of the J coupling over the frequency difference between A and B. In the case of an ALTADENA experiment, if the sample is slowly transferred toward the NMR magnet, polarization differences are definitely created and their amplitude can be larger than the amplitude of the longitudinal spin order. Some test experiments demonstrate the validity of the proposed theory.

International audience; Methionine sulfoxide reductases (Msr) reduce methionine sulfoxide (MetSO)-containing proteins, back to methionine (Met). MsrAs are stereospecific for the S epimer whereas MsrBs reduce the R epimer of MetSO. Although structurally unrelated, the Msrs characterized so far display a similar catalytic mechanism with formation of a sulfenic intermediate on the catalytic cysteine and a concomitant release of Met, followed by formation of at least one intramolecular disulfide bond (between the catalytic and a recycling cysteine), which is then reduced by thioredoxin. In the case of the MsrA from Escherichia coli, two disulfide bonds are formed, i.e. first between the catalytic Cys51 and the recycling Cys198 and then between Cys198 and the second recycling Cys206. Three crystal structures including E. coli and Mycobacterium tuberculosis MsrAs, which, for the latter, possesses only the unique recycling Cys198, have been solved so far. In these structures, the distances between the cysteine residues involved in the catalytic mechanism are too large to allow formation of the intramolecular disulfide bonds. Here structural and dynamical NMR studies of the reduced wild-type and the oxidized (Cys51-Cys198) forms of C86S/C206S MsrA from E. coli have been carried out. The mapping of MetSO substrate-bound C51A MsrA has also been performed. The data support (1) a conformational switch occurring subsequently to sulfenic acid formation and/or Met release that would be a prerequisite to form the Cys51-Cys198 bond and, (2) a high mobility of the C-terminal part of the Cys51-Cys198 oxidized form that would favor formation of the second Cys198-Cys206 disulfide bond.

International audience; Spectral density mapping at multiple NMR field strengths is probably the best method to describe the dynamical behavior of a protein in solution through the analysis of 15N heteronuclear relaxation parameters. Nevertheless, such analyses are scarcely reported in the literature, probably because this method is excessively demanding in spectrometer measuring time. Indeed, when using n different magnetic fields and assuming the validity of the high frequency approximation, the discrete sampling of the spectral density function with 2n + 1 points needs the measurement of 3n 15N heteronuclear relaxation measurements (n R 1, n R 2, and n15N{1H}NOEs). Based on further approximations, we proposed a new strategy that allows us to describe the spectral density with n + 2 points, with the measurement of a total of n + 2 heteronuclear relaxation parameters. Applied to the dynamics analysis of the protein p13 MTCP1 at three different NMR fields, this approach allowed us to divide by nearly a factor of two the total measuring time, without altering further results obtained by the “model free” analysis of the resulting spectral densities. Furthermore, simulations have shown that this strategy remains applicable to any low isotropically tumbling protein ($$\uptau_{c} > 3$$ ns), and is valid for the types of motion generally envisaged for proteins.

International audience; This study deals with a spin system constituted of three nonequivalent protons, two of them originating from para-hydrogen (p-H2) after a hydrogenation reaction carried out in the earth magnetic field. It is shown that three singlet states are created provided indirect (J) couplings exist between the three spins, implying hyperpolarization transfer toward the third spin. Upon insertion of the sample in the NMR (Nuclear Magnetic Resonance) high field magnet, the following events occur: (i) the longitudinal two-spin orders which are parts of the singlet states survive; (ii) the other two terms (of these singlet states) tend to be destroyed by magnetic field gradients but at the same time are partly converted into differences of longitudinal polarizations. Nuclear spin relaxation is studied by appropriate NMR measurements when evolution takes place in the high field magnet or in the earth field. In the former case, relaxation is classical although complicated by numerous relaxation rates associated with both longitudinal two-spin orders and longitudinal polarizations. In the latter case, an equilibration between the singlet states first occur, their disappearance being thereafter driven by relaxation rates which remain very small because of the absence of any dipolar contribution. Thus, even in the case of a three-spin system, long-lived states exist; this unexpected property could be very useful for many applications.

This study is an extension of a recent work (Canet D, Barthe P, Mutzenhardt P, Roumestand C. 2001. A comprehensive analysis of multifield 15N relaxation parameters in proteins: Determination of 15N chemical shift anisotropies. J Am Chem Soc 123:4567–4576) dealing with the exploitation of the three classical 15N relaxation parameters (the longitudinal relaxation rate, R1, the transverse relaxation rate, R2, and the 1H-15N cross-relaxation rate, σNH) measured at several magnetic fields (at least two) in uniformly 15N-labeled proteins (the chosen example is the C12A-p8MTCP1 protein whose relaxation parameters have been measured at five different fields). R2 is iteratively corrected for exchange, so that the three parameters provide consistently the NH dipolar and 15N csa (chemical shift anisotropy) contributions. The dipolar spectral densities are analyzed, residue by residue, according to the functional form (d for dipolar), where τeff is an (effective) correlation time associated with the slow motion(s) sensed by the NH vector at the level of the residue to which it belongs. In comparison with the Lipari-Szabo approach, the coefficient bd can be identified with 2KdτeffS2, S being the generalized order parameter and Kd reflecting the amplitude of the dipolar mechanism (which, besides constants, involves ); ad is associated with fast local motions. The csa spectral densities appearing in the expressions of R1 and R2 can be analyzed according to ; bcsa; is proportional to bd (the proportionality factor depending on molecular quantities) and can yield 15N csa values from the rNH value and from the knowledge of the angle between the two relevant tensors. Uncertainties are such that it is impossible (at least with the present data) to conclude about a possible variation of 15N csa values along the protein backbone. © 2005 Wiley Periodicals, Inc. Concepts Magn Reson Part A 24A: 1–9, 2005.

In part I of this article, it was shown that protein 15N relaxation data at different magnetic fields can be fitted according to a residue-specific correlation time τeff, which is associated with slow motions sensed by the considered residue. The present article is an attempt to justify the correlation time variations along the backbone by anisotropic overall motion. When an axial rotation-diffusion tensor is assumed, it is possible to derive simple expressions for relating τeff to τ⟂ and τ// (the two correlation times defining the rotation-diffusion tensor). A consistent picture is obtained for the most organized parts of the protein C12A-p8MTCP1 which, as in part I of this article, serves here as an example. By contrast, the model of rotation-diffusion fails for other less-organized parts of the protein, suggesting a combination of several slow or intermediate motions. © 2005 Wiley Periodicals, Inc. Concepts Magn Reson Part A 24A: 10–16, 2005.

A simple modification of the spin-state-selective excitation (S3E) pulse sequence element is proposed. The new sub-sequence, dubbed S2ED for spin-state echo differentiation, can be used in a very versatile way, in addition to the standard features of S3E. Experimental verifications were performed by editing α/β–HSQC–α/β spectra of nitrogen-15-labelled ubiquitin. Copyright © 2003 John Wiley & Sons, Ltd.

The cross-correlation relaxation rate constant between dipolar interaction and chemical shift anisotropy (interference term) is of considerable interest for structural and dynamical determinations. The present study focuses on methods related to its impact on longitudinal nuclear relaxation, and especially to a procedure based on the applications of a train of π pulses applied to both nuclei involved in such a process (Levitt MH, Di Bari L. Bull. Magn. Reson. 1994; 16: 94–114). The resulting steady state leads, in principle, to a straightforward determination of the relevant interference term. Simulations taking into account inhomogeneity of the radio-frequency (RF) field show that artifacts are responsible for instabilities which in practice preclude such a determination. An alternative method making use of a very limited number of RF pulses, combined with an appropriate filter, proves to be rather robust in view of achieving a proper measurement of the required interference term. Copyright © 2003 John Wiley & Sons, Ltd.

The assignments of 1H and 13C NMR spectra for three new triterpene saponins from Silene vulgaris (gypsogenin 3-O-glucuronide, quillaic acid 3-O-glucuronide, and gypsogenin 3-O-glycoside) are reported. In addition to 1D NMR methods, 2D NMR techniques (COSY, HSQC, HMBC, and HSQC-TOCSY) were used for the assignments. Copyright © 2002 John Wiley & Sons, Ltd.