1H NMR study of water relaxation and self-diffusion in human serum albumin aqueous solutions

Summary Water proton spin-lattice relaxation and self-diffusion in aqueous solutions of human serum albumin have been studied by¹H NMR as a function of the protein concentration. Spin-lattice relaxation data, which display a nonlinear behaviour with the protein concentration, could be fitted with a two-phase model taking into account the experimentally determined hydration (?bound?) water values. Despite a similar trend is registered for the water self-diffusion coefficient, such a model has been found unable to explain the related experimental data taken as a function of the biomolecule concentration. On the other hand, the solute-induced proton self-diffusion decrease could be satisfactorily interpreted by postulating an enhanced probability of hydrogen-bond formation occurring within the ?vicinal? water surrounding the biomolecules for several hydration shells. The consistency within the two models is discussed in connection with the magnetic interactions occurring within the solute-solvent systems.     

Unusual Increase of Apparent Transverse Relaxation Times in NMR Profiling of a Drying Polymer Solution

In this contribution, we present results from a nuclear magnetic resonance (NMR) profiling study of drying polymer solutions using polyvinylpyrrolidone (PVP) as a model system. The careful evaluation of the experimental data reveals a surprising increase in the apparent transverse relaxation time of the water phase of the drying solution over the first few hours of drying (where usually a decrease would be expected). By using additional information obtained from diffusion-weighted profiles, we can explain this finding as the result of a strong reduction of the diffusion effect on the apparent relaxation time during this time interval. Understanding this effect provides a better basis for making optimal use of profiling NMR in drying studies and avoids possible misinterpretation on the data.     

Porous structure of membranes of an acrylonitrile copolymer. Porosity, 1H-NMR permeability

Nanoporous polymer membranes (porosity ) used for dialysis are studied from NMR relaxation times of water confined in the pore space. Fast interpore water diffusion is observed. Two structural parameters are evidenced: i) a reduced NMR relaxation time, , which reflects the width of the pore-size distribution; ii) the average polymer-grain size of the solid matrix deduced from NMR experiments performed on membranes partially filled by water. A relation is found between the ratio , where k is the permeability to water and the porosity. This relation is in qualitative agreement with numerical simulations reported in the literature on low-porosity systems and with experimental results obtained for sedimentary rocks and for fused glass model systems. It supports the idea that is the relevant structural parameter to describe convective transport in a wide class of porous systems. Received 8 July 1999     

DOUBLESLICING: a non-iterative single profile multi-exponential curve resolution procedure. Application to time-domain NMR transverse relaxation data

A new non-iterative curve resolution technique for resolving single decay profiles is proposed. The new technique, called DoubleSlicing, is based on the Decra (Direct Exponential Curve Resolution Algorithm) principle. While the original Decra was designed to resolve several decay curves simultaneously and thus fitting common pure exponentials, DoubleSlicing can resolve single decay profiles by a simple double data transformation followed by an analytical and unique three-way decomposition. The new approach is successfully demonstrated on experimental NMR CPMG relaxation data, measured on combinations of unmixed paramagnetic CuSO(4) solutions. Decay signals of the water component were acquired following an innovative experimental design that ensured no interaction between the components present in each sample under observation. DoubleSlicing proved to be accurate in estimating relaxation times differing in one order of magnitude (range: 19.6-159.4ms). Its performance was comparable to discrete exponential fitting with the advantage of being much faster - in terms of computation time, DoubleSlicing outperformed exponential fitting by a factor of four.     

NMR relaxation of protons in tissues and other macromolecular water solutions

Nuclear magnetic resonance (NMR) longitudinal (T₁) and transverse (T₂) relaxation parameters have been evaluated for protein solutions, cellular suspensions and tissues using both data from our laboratory and the extensive literature. It is found that this data can be generalized and explained in terms of three water phases: free water, hydration water, and crystalline water. The proposed model which we refer to as the FPD model differs from similar models in that it assumes that free and hydration water are two phases with distinct relaxation times but that T₁ = T₂ in each phase. In addition there is a single correlation time for each rather than a distribution as assumed in most other models. Longitudinal decay is predicted to be single exponent in character resulting from a fast exchange between the free and hydration compartments. Transverse decay is predicted to be multiphasic with crystalline (T₂ 10 μsec), hydration (T₂ 10 sec) and free (T₂ 100 sec) water normally visible. The observed or effective transverse relaxation times for both the hydration and free water phases are greatly affected by the crystalline phase and are much shorter than the inherent relaxation times.     

Dynamics of paramagnetic agents by off-resonance rotating frame technique

Off-resonance rotating frame technique offers a novel tool to explore the dynamics of paramagnetic agents at high magnetic fields (B0 > 3T). Based on the effect of paramagnetic relaxation enhancement in the off-resonance rotating frame, a new method is described here for determining the dynamics of paramagnetic ion chelates from the residual z-magnetizations of water protons. In this method, the dynamics of the chelates are identified by the difference magnetization profiles, which are the subtraction of the residual z-magnetization as a function of frequency offset obtained at two sets of RF amplitude omega(1) and pulse duration tau. The choices of omega(1) and tau are guided by a 2-D magnetization map that is created numerically by plotting the residual z-magnetization as a function of effective field angle theta and off-resonance pulse duration tau. From the region of magnetization map that is the most sensitive to the alteration of the paramagnetic relaxation enhancement efficiency R(1rho)/R1, the ratio of the off-resonance rotating frame relaxation rate constant R(1rho) verse the laboratory frame relaxation rate constant R(1), three types of difference magnetization profiles can be generated. The magnetization map and the difference magnetization profiles are correlated with the rotational correlation time tauR of Gd-DTPA through numerical simulations, and further validated by the experimental data for a series of macromolecule conjugated Gd-DTPA in aqueous solutions. Effects of hydration water number q, diffusion coefficient D, magnetic field strength B0 and multiple rotational correlation times are explored with the simulations of the magnetization map. This method not only provides a simple and reliable approach to determine the dynamics of paramagnetic labeling of molecular/cellular events at high magnetic fields, but also a new strategy for spectral editing in NMR/MRI based on the dynamics of paramagnetic labeling in vivo.     

NMR 1D-imaging of water infiltration into mesoporous matrices

It is shown that coupling nuclear magnetic resonance (NMR) 1D-imaging with the measure of NMR relaxation times and self-diffusion coefficients can be a very powerful approach to investigate fluid infiltration into porous media. Such an experimental design was used to study the very slow seeping of pure water into hydrophobic materials. We consider here three model samples of nuclear waste conditioning matrices which consist in a dispersion of NaNO₃ (highly soluble) and/or BaSO₄ (poorly soluble) salt grains embedded in a bitumen matrix. Beyond studying the moisture progression according to the sample depth, we analyze the water NMR relaxation times and self-diffusion coefficients along its 1D-concentration profile to obtain spatially resolved information on the solution properties and on the porous structure at different scales. It is also shown that, when the relaxation or self-diffusion properties are multimodal, the 1D-profile of each water population is recovered. Three main levels of information were disclosed along the depth-profiles. They concern (i) the water uptake kinetics, (ii) the salinity and the molecular dynamics of the infiltrated solutions and (iii) the microstructure of the water-filled porosities: open networks coexisting with closed pores. All these findings were fully validated and enriched by NMR cryoporometry experiments and by performing environmental scanning electronic microscopy observations. Surprisingly, results clearly show that insoluble salts enhance the water progression and thereby increase the capability of the material to uptake water.     

用异核多维NMR技术研究蛋白质动力学

蛋白质在溶液中的三维空间结构、动力学与蛋白质生物功能的关系是在分子水平上理解生命现象的重要基础. NMR技术在研究蛋白质动力学方面具有独特的优势,所能表征的运动过程相关时间尺度很广. 文章综述了异核多维NMR技术研究蛋白质动力学的实验技术和理论方法,介绍了描述蛋白质动力学的内运动参量的意义和Model-Free 方法,并举例说明¹⁵N弛豫测量实验被用于研究蛋白质及其与配体复合物的动力学.      

Dynamics of conformations,hydrogen bonds and translational diffusion of poly(methacrylic acid) in aqueous solution and the concentration transition in MD simulations

The dynamic behaviour of chain conformations, hydrogen bonds and translational diffusion of aqueous poly(methacrylic acid) (PMA) solution as a function of polymer volume fraction Φ_p across dilute to concentrated regimes inclusive of the pure polymer amorphous state was studied by molecular dynamics simulations. The behaviour of the relaxation time (τ) of the backbone dihedral angle auto-correlation function (ACF) reveals slower relaxation at higher level of polymer concentration and the existence of a concentration-driven relaxation transition for the aqueous polymer solution which occurs in the polymer volume fraction range, specifically 54% < Φ_p < 82% for this system. The relaxation constant τ for backbone dihedral angle exhibits a linear variation with Φ_p, indicating a first-order kinetic transition. The intermittent ACF for decay of the H-bond correlation shows that H-bonds among water molecules relax faster than those of the PMA–PMA and PMA–water type. The relaxation rate of PMA–water H-bonds shows a decrease up to Φ_p = 72% and becomes faster at Φ_p = 82% due to the confining influence of neighbouring PMA chains. PMA–water and water–water H-bond dynamics show transitions around Φ_p = 72% PMA. With increase in Φ_p PMA diffusion coefficient decreases exponentially and water diffusion coefficient decreases linearly, in agreement with experimental observations using fluorescence and nuclear magnetic resonance (NMR) spectroscopic studies.     

A faster way to characterize by triple-quantum-filtered (17)O NMR water molecules strongly bound to macromolecules in solution

The simultaneous use of transverse and longitudinal relaxation rates, together with a transverse triple-quantum-filtering NMR sequence, was estimated for the adequate characterization of (17)O-water relaxation behavior in protein solutions. A complementary contribution to transverse relaxation was found, which was interpreted as chemical exchange of (17)O-water between different sites of the proteins. This contribution was estimated via calibration measurements. Then, for other similar samples, faster experiments could be performed. The analysis of the results obtained in this way gave adequate values of the relaxation rate of water in fast motion, of the fraction of water in slow motion, and of its correlation time. Hence, it permitted the complete characterization of the sample in a reasonable experimental time.     

On a hydration model utilized in the discussion of dielectric spectra of aqueous solutions

A solution model is discussed which allows the microwave part of the permittivity spectrum of aqueous solutions to be related to characteristics of the hydration water. The parameters, which can be derived from measured dielectric spectra thereby are the hydration water relaxation time, the number of hydration water molecules per molecule of solute, the static orientational polarizability of the hydration water, and a quantity, which refers to the distribution of hydration water relaxation times. The (continuum) model, appropriate for solutions of (nearly) spherically shaped solute particles, has regard to internal electric fields resulting from polarization charges at interfaces. Possible errors in the parameter values are indicated, which may arise if the internal fields are only incompletely taken into account. Previously measured spectra for a series of aqueous solutions of 1,4-diazabicyclo[2,2,2]octane have been evaluated on the basis of the present model. The results for these (favourable) solutions are presented to show, that the found dependence of the parameter values on solute concentration is consistent with the idea of the proposed hydration model.     

On the strong field dependence and nonlinear response to gadolinium contrast agent of proton transverse relaxation rates in dairy cream

Dairy cream, as a suspension of lipid droplets in water, is a potentially useful magnetic resonance imaging (MRI) phantom material and an interesting material for studying fundamental relaxation mechanisms. Here we report a strong increase in the transverse relaxation rates with field strength for both the water and lipid protons in dairy cream. Also, studies at 4.7 T reveal a nonlinear response of transverse relaxation rates with increasing concentration of a common gadolinium (Gd)-based contrast agent, including an initial decrease of water relaxation rates as measured with Hahn spin echoes at the lower Gd concentrations. The results are treated within the framework of a model in which the magnetic susceptibility difference between the lipid droplets and the aqueous phase plays the prominent role for transverse relaxation. Second-order polynomial fits of the water proton transverse relaxation rate dependence on field strength and on Gd concentration at 4.7 T provided experimental parameters from which model parameters are extracted and compared with expectations available from the literature.     

The matrix formalism for generalised gradients with time-varying orientation in diffusion NMR

Protein backbone 15N NMR spin relaxation rates are useful in characterizing the protein dynamics and structures. To observe the protein nuclear-spin resonances a pulse sequence has to include a water suppression scheme. There are two commonly employed methods, saturating or dephasing the water spins with pulse field gradients and keeping them unperturbed with flip-back pulses. Here different water suppression methods were incorporated into pulse sequences to measure 15N longitudinal T1 and transversal rotating-frame T1ρ spin relaxation. Unexpectedly the 15N T1 relaxation time constants varied significantly with the choice of water suppression method. For a 25-kDa Escherichiacoli. glutamine binding protein (GlnBP) the T1 values acquired with the pulse sequence containing a water dephasing gradient are on average 20% longer than the ones obtained using a pulse sequence containing the water flip-back pulse. In contrast the two T1ρ data sets are correlated without an apparent offset. The average T1 difference was reduced to 12% when the experimental recycle delay was doubled, while the average T1 values from the flip-back measurements were nearly unchanged. Analysis of spectral signal to noise ratios (s/n) showed the apparent slower 15N relaxation obtained with the water dephasing experiment originated from the differences in 1HN recovery for each relaxation time point. This in turn offset signal reduction from 15N relaxation decay. The artifact becomes noticeable when the measured 15N relaxation time constant is comparable to recycle delay, e.g., the 15N T1 of medium to large proteins. The 15N relaxation rates measured with either water suppression schemes yield reasonable fits to the structure. However, data from the saturated scheme results in significantly lower Model-Free order parameters (=0.81) than the non-saturated ones (=0.88), indicating such order parameters may be previously underestimated.     

Understanding Overhauser Dynamic Nuclear Polarisation through NMR relaxometry

Overhauser dynamic nuclear polarisation (DNP) represents a potentially outstanding tool to increase the sensitivity of solution and solid state NMR experiments, as well as of magnetic resonance imaging. DNP signal enhancements are strongly linked to the spin relaxation properties of the system under investigation, which must contain a paramagnetic molecule used as DNP polariser. In turn, nuclear spin relaxation can be monitored through NMR relaxometry, which reports on the field dependence of the nuclear relaxation rates, opening a route to understand the physical processes at the origin of the Overhauser DNP in solution. The contributions of dipole–dipole and Fermi-contact interactions to paramagnetic relaxation are here described and shown to be responsible to both the relaxometry profiles and the DNP enhancements, so that the experimental access to the former can allow for predictions of the latter.     

Molecular dynamics of n-dodecylammonium chloride in aqueous solutions investigated by 2H NMR and 1H NMR relaxometry

Molecular dynamics in n-dodecylammonium chloride/water solutions for concentrations of 34 and 45 wt% was studied by 2H NMR and by 1H NMR dispersion of spin-lattice relaxation in the 2 kHz-90 MHz frequency range. The system exhibits a number of lyotropic liquid crystalline phases, which differ in symmetry and involve motions characterized by a wide frequency scale. The analysis of 2H NMR lineshapes of selectively deuterated DDACl molecules gave us an evidence for local trans-gauche conformational changes in the chains, whereas the dispersion of spin-lattice relaxation times T1 explored by fast field cycling method revealed fast local motions, translational diffusion and collective molecular dynamics of the chains. In particular, we have found that the order director fluctuation mechanism in smectic and nematic phases dominates spin-lattice relaxation below 1 MHz and that local motions and translational diffusion are responsible for the spin-lattice relaxation in the higher Larmor frequency range.     

A new approach to visualizing spectral density functions and deriving motional correlation time distributions: applications to an alpha-helix-forming peptide and to a well-folded protein

A new approach to visualizing spectral densities and analyzing NMR relaxation data has been developed. By plotting the spectral density function, J(omega), as F(omega)=2 omega J(omega) on the log-log scale, the distribution of motional correlation times can be easily visualized. F(omega) is calculated from experimental data using a multi-Lorentzian expansion that is insensitive to the number of Lorentzians used and allows contributions from overall tumbling and internal motions to be separated without explicitly determining values for correlation times and their weighting coefficients. To demonstrate the approach, (15)N and (13)C NMR relaxation data have been analyzed for backbone NH and C(alpha)H groups in an alpha-helix-forming peptide 17mer and in a well-folded 138-residue protein, and the functions F(omega) have been calculated and deconvoluted for contributions from overall tumbling and internal motions. Overall tumbling correlation time distribution maxima yield essentially the same overall correlation times obtained using the Lipari-Szabo model and other standard NMR relaxation data analyses. Internal motional correlational times for NH and C(alpha)H bond motions fall in the range from 100 ps to about 1 ns. Slower overall molecular tumbling leads to better separation of internal motional correlation time distributions from those of overall tumbling. The usefulness of the approach rests in its ability to visualize spectral densities and to define and separate frequency distributions for molecular motions.     

Structural relaxation of polymers at the glass transition: conformational memory in poly(n-alkylmethacrylates)

Conformational memory is introduced as a new aspect of structural relaxation of polymers. In poly(n-alkylmethacrylates) extended backbone chain conformations are identified by advanced NMR techniques as the molecular units involved in structural relaxation. They retain conformational memory over many steps of restricted axial chain motion. Randomization of conformation and isotropization of backbone orientation occur on the same time scale, yet much slower than the slowest relaxation process identified so far. Behavior typical of fragile glass formers is found for this new process of chain relaxation.     

Proton NMR and susceptibility measurements on the magnetic core of ferritin

We report an investigation of the magnetic core of the biomolecule ferritin by means of proton nuclear magnetic resonance (NMR) and relaxation, magnetic susceptibility and scanning electron microscope (SEM) measurements. SEM images show that the outer protein shell is taken out completely by an appropriate chemical treatment and indicate particle sizes ranging from 10² to 10⁴ nm. Susceptibility measurements show a maximum in the zero-field-cooled data which is strongly field-dependent and can be ascribed to superparamagnetic behavior, whereas the hysteresis curve is different from normal ferritin. Proton NMR and spin-lattice relaxation data as a function of temperature at 4.7 T suggest the presence of an antiferromagnetic transition around 100 K.     

On-line Surface Area Measurement of Concentrated Slurries using low field spin-lattice relaxation NMR

A method for the rapid on-line determination of surface area and solids content in flowing concentrated slurries using low field NMR spin-lattice relaxation measurements has been developed and demonstrated. The relationship between flow and spin-lattice relaxation time (T₁) of protons in water at 20 MHz was examined using aqueous copper sulfate solutions. The ability to measure surface area and solids concentration in both stagnant (stopped flow) and flowing systems via NMR was demonstrated using several different concentrated aqueous titania and glass slurries (20 to 80 weight percent) for which the dried powder surface area was previously determined via nitrogen adsorption/BET analysis and the solids content determined gravimetrically. Surface areas were also calculated from particle size analysis and found to vary by up to an order of magnitude from the adsorption and NMR results.