Nuclear magnetic resonance transverse relaxation in muscle water.

The origin of the nonexponentiality of proton spin echoes of skeletal muscle has been carefully examined. It is shown that the slowly decaying part of the proton spin echoes is not due to extracellular water. First, for muscle from mice with in vivo deuteration, the deuteron spin echoes were also nonexponential, but the slowly decaying part had a larger weighing factor. Second, for glycerinated muscle in which cell membranes were disrupted, the proton spin echoes were similar to those in intact muscle. Third, the nonexponentiality of the proton spin echoes in intact muscle increased when postmortem rigor set in. Finally, when the lifetimes of extracellular water and intracellular water were taken into account in the exchange, it was found that the two types of water would not give two resolvable exponentials with the observed decay constants. It is suggested that the unusually short T2's and the nonexponential character of the spin echoes of proton and deuteron in muscle water are mainly due to hydrogen exchange between water and functional groups in the protein filaments. These groups have large dipolar or quadrupolar splittings, and undergo hydrogen exchange with water at intermediate rates. The exchange processes and their effects on the spin echoes are pH-dependent. The dependence of transverse relaxation of pH was observed in glycerinated rabbit psoas muscle fibers.     

Study of anisotropy in nuclear magnetic resonance relaxation times of water protons in skeletal muscle.

The anisotropy of the spin-lattice relaxation time (T1) and the spin-spin relaxation times (T2) of water protons in skeletal muscle tissue have been studied by the spin-echo technique. Both T1 and T2 have been measured for the water protons of the tibialis anterior muscle of mature male rats for theta = 0, 55, and 90 degrees, where theta is the orientation of the muscle fiber with respect to the static field. The anisotropy in T1 and T2 has been measured at temperatures of 28, -5 and -10 degrees C. No significant anisotropy was observed in the T1 of the tissue water, while an average anisotropy of approximately 5% was observed in T2 at room temperature. The average anisotropy of T2 at -5 and -10 degrees C was found to be approximately 2 and 1.3%, respectively.     

Interstitial sodium nuclear magnetic resonance relaxation times in perfused hearts.

Separation of intracellular and extracellular sodium nuclear magnetic resonance (NMR) signals would enable nondestructive monitoring of intracellular sodium. It has been proposed that differences between the relaxation times of intracellular and extracellular sodium be used either directly or indirectly to separate the signal from each compartment. However, whereas intracellular sodium relaxation times have been characterized for some systems, these times were unknown for interstitial sodium. In this study, the interstitial sodium NMR relaxation times have been measured in perfused frog and rat hearts under control conditions. This was achieved by eliminating the NMR signal from the extracardiac (perfusate) sodium, and then quantifying the remaining cardiac signal. The intracellular signal was measured to be 8% (frog) or 22% (rat) of the cardiac signal and its subtraction was found to have a negligible effect on the cardiac relaxation times. Therefore this cardiac signal is considered to provide a good estimate of interstitial relaxation behavior. For perfused frog (rat) hearts under control conditions, this signal was found to have a T1 of 31.6 +/- 3.0 ms (27.3 +/- 1.6 ms) and a biexponential T2 of 1.9 +/- 1.0 ms (2.1 +/- 0.3 ms) and 25.2 +/- 1.3 ms (26.3 +/- 3.2 ms). Due to the methods used to separate cardiac signal from perfusate signal, it is possible that this characterized only a part of the signal from the interstitium. The short T2 component attributable to the interstitial signal indicates that separation of the NMR signals from each compartment on the basis of relaxation times alone may be difficult.     

Nuclear magnetic resonance study of spin relaxation and magnetic field gradients in maple leaves.

1H Nuclear magnetic resonance techniques were used to measure the distributions of spin-spin relaxation times, T2, and of magnetic field gradients in both the chloroplast and nonchloroplast water compartments of maple leaves (Acer platanoides). Results showed that encounters between water molecules and membranes inside chloroplasts provide an inefficient relaxation mechanism; i.e., chloroplast membranes interact weakly with water molecules. Gradient measurements indirectly measured the sizes of chloroplasts by showing that water in the chloroplasts is confined to small compartments a few microns in diameter. A comparison between measured gradients and gradients calculated for a model leaf indicated that chloroplasts are somewhat more likely to occupy positions along cell walls adjacent to air spaces, but also they may be found in the interiors of cells.     

Effects of temperature and pH on the water exchange through erythrocyte membranes: Nuclear magnetic resonance studies

Summary The temperature and pH dependence of water exchange has been studied on isolated erythrocytes suspended in isotonic buffered solutions. At pH 7.4 a break in the Arrhenius plot of water exchange time at around 26°C was found. The mean value of the apparent activation energy of the water exchange time at temperatures higher than that of the discontinuity was 5.7 kcal/mole (±0.4); at lower temperatures the values of the apparent activation energy were below 1.4 kcal/mole. The pH dependence of water exchange time of isolated erythrocytes revealed a marked increase of the water exchange time values in the acid range of pH; a much smaller variation of the same parameter occurs between pH 7.0 and 8.0. These finding could be correlated with other processes involving erythrocyte membranes that showed similar pH and temperature dependence and were considered to indicate state transitions in the membranes. It is suggested that the temperature and pH effects on water diffusion indicate that conformational changes and cooperative effects are implicated in the mechanism of this transport process.Institute for Isotopic and Molecular Technology.     

Nuclear magnetic resonance relaxation time and self-diffusion coefficient measurements of water in frog ovarian eggs (Rana pipiens)

Self-diffusion coefficient measurements of water in untreated ovarian eggs of Rana pipiens using nuclear magnetic resonance indicate that cytoplasmic water has reduced translational mobility compared with pure water. Using a simple two-state model, we find that ～67% is “relatively immobile.” Consideration of the nuclear magnetic resonance spin-lattice and spin-spin relaxation times indicates that the decreased mobility can largely be ascribed to hydration. Our value for the self-diffusion coefficient (6.8 × 10^?6 cm²/sec) is lower than those reported by other investigators using isotopic water exchange techniques on frog eggs chemically treated to remove the membrane. However, the results reported here are in agreement with unpublished data on untreated frog eggs implying that chemical treatment has modified the cytoplasm in some manner.     

Nuclear magnetic resonance relaxation in cross-linked lysozyme crystals: an isotope dilution experiment.

Nuclear magnetic resonance (nmr) relaxation times are measured for water protons in cross-linked lysozyme crystals below the freezing event as a function of the mole fraction of protons in the water phase. Proton longitudinal nmr relaxation in these samples is nonexponential and the slow longitudinal relaxation component becomes slower linearly with decreasing proton mole fraction in the water. The data are analyzed using a cross relaxation model that eliminates the necessity of postulating long residence times for water molecules in the domain of the protein. The observed isotope dilution behavior is consistent with the cross relaxation model. The deuterium nmr relaxation is also reported for deuterium oxide in the cross-linked protein crystal sample below the freezing event and the relaxation is shown to be accurately exponential.     

Nuclear magnetic resonance studies of amino acids and proteins. Rotational correlation times of proteins by deuterium nuclear magnetic resonance spectroscopy

We show that measurement of the spin-lattice (T1) and spin-spin (T2) relaxation times (or line widths) of irrotationally bound 2H nuclei in macromolecules undergoing isotropic rotational motion outside of the extreme narrowing limit (i.e., for the case omega 02 tau R2 much greater than 1) permits determination of both the rotational correlation time (tau R) of the macromolecule and the electric quadrupole coupling constant (e2qQ/h) of the 2H label. The technique has the advantage over 13C nuclear magnetic resonance (NMR) that no assumptions about bond lengths (which appear to the sixth power in 13C relaxation studies) or relaxation mechanisms need to be made, since relaxation will always be quadrupolar, even for aromatic residues at high field. Asymmetry parameter (eta) uncertainties are shown to cause negligible effects on tau R determinations, and in any case it is shown that both e2qQ/h and eta may readily be determined in separate solid-state experiments. By way of example, we report 2H NMR results on aqueous lysozyme (EC 3.2.1.17) at 5.2 and 8.5 T (corresponding to 2H-resonance frequencies of 34 and 55 MHz). Interpretation of the results in terms of the isotropic rigid-rotor model yields e2qQ/h values of approximately equal to 170 or approximately equal to 190 kHz, respectively, for the imidazolium and free-base forms of [epsilon 1-2H] His-15 lysozyme in solution, in excellent agreement with e2qQ/h values of approximately 167 and approximately 190 kHz obtained for the free amino acids in the solid state. In principle, the method may in suitable cases permit comparison between the dynamic structures of proteins in solution and in the crystalline solid state.     

Nuclear magnetic resonance of water in cold acclimating red osier dogwood stem

The pulsed and continuous-wave nuclear magnetic resonance of water in cold-acclimating red osier dogwood (Cornus stolonifera Michx) stem showed reduced relaxation times and increased line width. The reduction of relaxation times suggests an over-all restriction in the motional characteristics of the water. The increased line width is not related to a molecular property of the water, but is useful in estimating the initiation of cold acclimation. Biphasic relaxation characteristics may be related to partitioning of the water at the cellular level. The liquid water content of the stem was a weak function of temperature between −25 and −55 C, corresponding to approximately 0.15 gram of water per gram of dry stem. The quantity of unfrozen water at subfreezing temperatures was not strongly dependent on the degree of cold acclimation. It is concluded that the ability of dogwood to survive low temperatures depends on its ability to tolerate diminished quantities of liquid water.     

Nuclear magnetic resonance investigation of the state of water in human hair

Measurements have been made of the nuclear magnetic relaxation times T₁ and T₂ of the protons of water in hair. These are interpreted as showing that water molecules in hair exist in a continuous range of environments with a wide spread of rates of molecular rotation. Even at high water contents most of the water molecules are much less mobile than molecules in bulk water. The term “mobility” is given a quantitative meaning.     

Nuclear magnetic resonance and protein structure

NMR provides a wealth of structural information about proteins in solution, but does not, by itself, permit an unambiguous determination of a unique structure. A rigorous interpretation of NMR data to obtain the entire family of structures compatible with a given data set requires extensive, systematic and unbiased sampling of the conformational space of the polypeptide chain. Methods of sampling based on the exclusion paradigm--i. e. those that generate structures, check constraints and accept or reject members of the family on that basis, avoid the problem of generating erroneous structures by converging on local minima, which is a common pitfall of methods based on the optimization paradigm. Their much higher computational cost can be reduced by solving the structure in stages, using abstract representations of partial structures, and guiding the computation by control heuristics. The heuristic refinement method developed at Stanford and encoded in the expert system PROTEAN yields more or less extensive families of structures, depending on the size of the NMR data set, and defines the "allowed volume" in which each atom (or other substructure) may lie, with all experimental constraints satisfied. The allowed volume is a measure of the uncertainty of our knowledge of the structure, to which both the limitations of the data and the uncertainty of position resulting from molecular motion may contribute. Prediction of the experimental NMR spectra by solving the generalized Bloch equations (or the Redfield density matrix) for the protein, using atomic coordinates that lie within the allowed atomic volume, provides the final test for the correctness of the proposed structure.(ABSTRACT TRUNCATED AT 250 WORDS)     

Nuclear magnetic resonance of lipid A--the influence of solvents on spin relaxation and spectral quality

Nuclear magnetic resonance (NMR) spectroscopy of lipid A is limited by rapid transversal relaxation and subsequent line broadening caused by the tendency of these glycolipids to form aggregates in all solvents. To examine the influence of solvents on NMR spectra, hexa-acyl lipid A from Escherichia coli F515 was investigated. Line widths at half height, longitudinal relaxation times, and transversal relaxation times were measured in different solvents, lipid A concentrations, and temperatures. Chloroform-d, dioxane-d(8), and pyridine-d(5) each mixed with 25% methanol-d(4) as well as sole DMSO-d(6) and 0.1M triethylamine-d(15) (TEA-d(15)) in D(2)O caused good spectral resolutions and allowed structure analysis. ROESY and HMBC spectra gave an insight into the influence of transversal relaxation times on spectral quality in two-dimensional spectra. Solvent depending differences of interglycosidic NOEs indicated dissimilarities of the conformations in the interglycosidic linkage and allowed conclusions about the lipid A solution state.     

Nuclear magnetic spin-lattice relaxation of water protons caused by metal cage compounds.

The nuclear magnetic spin-lattice relaxation rates of water protons are reported for solutions of manganese(II), copper(II), and chromium(III) cage complexes of the sarcophagine type. As simple aqueous solutions, the complexes are only modest magnetic relaxation agents, presumably because they lack protons on atoms in the first-coordination-sphere protons that are sufficiently labile to mix the large relaxation rate at the metal complex with that of the bulk solvent. The relaxation is approximately modeled using spectral density functions derived for translational diffusion of the interacting dipole moments with the modification that the electron spin relaxation rate is directly included as a contribution to the correlation time. In all cases studied, the electron spin relaxation rate is sufficiently large that it contributes directly to the water-proton spin relaxation process. The poor relaxation efficiency of the cage compound may, however, be improved dramatically by binding the complex to a protein. The efficiency is improved even further if the rotational motion of the protein is reduced drastically by an intermolecular cross-linking reaction. The relaxation efficiency of the cross-linked protein-cage complexes rivals that of the best first-coordination-sphere relaxation agents like [Gd(DTPA)(H2O)]2- and [Gd(DOTA)(H2O)]-.