Characterization of the backbone dynamics of an anti-digoxin antibody VL domain by inverse detected 1H–15N NMR: Comparisons with X-ray data for the Fab

The dynamic behavior of the polypeptide backbone of a recombinant anti-digoxin antibody V_L domain has been characterized by measurements of ¹⁵N T₁ and T₂ relaxation times, ¹H–¹⁵N NOE values, and ¹H–²H exchange rates. These data were acquired with 2D inverse detected heteronuclear ¹H–¹⁵N NMR methods. The relaxation data are interpreted in terms of model free spectral density functions and exchange contributions to transverse relaxation rates R₂ (= 1/T₂). All characterized residues display low-amplitude picosecond timescale librational motions. Fifteen residues undergo conformational changes on the nanosecond timescale, and 24 residues have significant R₂ exchange contributions, which reflect motions on the microsecond to millisecond timescale. For several residues, microsecond to millisecond motions of nearby aromatic rings are postulated to account for some or all of their observed R₂ exchange contributions. The measured ¹H–²H exchange rates are correlated with hydrogen bonding patterns and distances from the solvent accessible surface. The degree of local flexibility indicated by the NMR measurements is compared to crystallographic B-factors derived from X-ray analyses of the native Fab and the Fab/digoxin complex. In general, both the NMR and X-ray data indicate enhanced flexibility in the turns, hypervariable loops, and portions of β-strands A, B, and G. However, on a residue-specific level, correlations among the various NMR data, and between the NMR and X-ray data, are often absent. This is attributed to the different dynamic processes and environments that influence the various observables. The combined data indicate that certain regions of the V_L domain, including the three hypervariable loops, undergo dynamic changes upon V_L:V_H association and/ or complexation with digoxin. Overall, the 26–10 V_L domain exhibits relatively low flexibility on the ps–ns timescale. The possible functional consequences of this result are considered. © 1993 Wiley-Liss, Inc.     

Structural,photoluminescence, and optical characteristics of a Sm3+-doped lead borate–strontium–tungsten glass system: Evaluation of Judd–Ofelt intensity parameters and radiative properties

In this study, the melt quenching approach is used to synthesize a lead borate–strontium-based glass system doped with samarium ions. Modifications in the glass network structure arising from the addition of various concentrations of Sm³⁺ ions were investigated via Fourier transform infrared (FTIR) spectroscopy. FTIR analysis revealed B–O–B bridges, BO₃, and BO₄ units are present. UV–vis–NIR spectroscopic measurement was performed to study the optical absorption spectra. Optical constants such as optical bandgap energies, refractive indices, and other related parameters were evaluated. The lifetime fluorescence decay was measured and ranged between 1.04 and 1.88 ns. The photoluminescence spectra in the range 500–750 nm revealed four transitions from the ground state ⁶G_5/2 to the excited states ⁶H_5/2, ⁶H_7/2, ⁶H_9/2 and ⁶H_11/2 and J–O theory was utilized to study these optical transitions for Sm³⁺ ions. Calculations of the oscillator strengths and J–O intensity parameters were performed and the obtained J–O parameters followed the sequence Ω₄ > Ω₆ > Ω₂. The ratio O/R indicated a high lattice asymmetry around the samarium ions. The values of lifetimes and branching ratios for the fabricated samples emphasized their suitability to be used in laser applications. The current glass samples are good candidates for orange and red emission devices.     

Intracellular water in Artemia cysts (brine shrimp): Investigations by deuterium and oxygen-17 nuclear magnetic resonance

The dormant cysts of Artemia undergo cycles of hydration-dehydration without losing viability. Therefore, Artemia cysts serve as an excellent intact cellular system for studying the dynamics of water-protein interactions as a function of hydration. Deuterium spin-lattice (T₁) and spin-spin (T₂) relaxation times of water in cysts hydrated with D₂O have been measured for hydrations between 1.5 and 0.1 g of D₂O per gram of dry solids. When the relaxation rates (I/T₁, I/T₂) of ²H and ¹⁷O are plotted as a function of the reciprocal of hydration (1/H), an abrupt change in slope is observed near 0.6 g of D₂O (or H₂ ¹⁷O)/gram of dry solids, the hydration at which conventional metabolism is activated in this system. The results have been discussed in terms of the two-site and multisite exchange models for the water-protein interaction as well as protein dynamics models. The ²H and ¹⁷O relaxation rates as a function of hydration show striking similarities to those observed for anisotropic motion of water molecules in protein crystals.

It is suggested here that although the simple two-site exchange model or n-site exchange model could be used to explain our data at high hydration levels, such models are not adequate at low hydration levels (<0.6 g H₂O/g) where several complex interactions between water and proteins play a predominant role in the relaxation of water nuclei. We further suggest that the abrupt change in the slope of I/T₁ as a function of hydration in the vicinity of 0.6 g H₂O/g is due to a change in water-protein interactions resulting from a variation in the dynamics of protein motion.

     

An exploratory scandium-45 NMR study into the complexation of alanine and oligopeptides

The 87.5 MHz ⁴⁵Sc NMR spectrum of 0.025 M aqueous Sc(NO₃)₃ exhibits two resonance signals, separated by ca. 25 ppm, attributable to [Sc(H₂O)₆]³⁺ and [Sc(H₂O)₅OH]²⁺. Acidification leads to a single, comparatively sharp line (W_1/2 = 160 Hz) for the hexaqua complex, the temperature dependence (temperature gradient = 0.076 ppm/deg) of which indicates that relaxation is dominated by the quadrupole mechanism. Addition of α-alanine gives rise to an additional broad signal at ca. +70 ppm (relative to [Sc(H₂O)₆]³⁺), which is assigned to a carboxylato complex [Sc(H₂O)_6−n(ala)_n]³⁺ or [Sc(H₂O)_5−nOH- (ala)_n]²⁺ (1 < n < 2). At ambient temperatures, these species are in slow exchange with the hexaqua and pentaqua-hydroxo complex, progressing through medium towards fast exchange as the temperature increases, and giving rise to an exchange contribution to relaxation. W_1/2 becomes a measure for the stability of the complexes, which increases in the order ala < (ala)₄ ∼ (ala)₂ < ala-val-leu. The pronounced stability of the latter is due to the formation of a chelate-five ring structure (participation of the NH- function of the peptide bond in coordination to Sc³⁺). 1 M aqueous ScCl₃ probably contains the two species [Sc(H₂O)₆]³⁺ and [Sc(H₂O)₅Cl]²⁺, separated by 33 ppm.     

Mechanistic Insights of Zn2+ Storage in Sodium Vanadates

Rechargeable aqueous zinc‐ion batteries (ZIBs) with high safety and low‐cost are highly desirable for grid‐scale energy storage, yet the energy storage mechanisms in the current cathode materials are still complicated and unclear. Hence, several sodium vanadates with NaV₃O₈‐type layered structure (e.g., Na₅V₁₂O₃₂ and HNaV₆O₁₆·4H₂O) and β‐Na_0.33V₂O₅‐type tunneled structure (e.g., Na_0.76V₆O₁₅) are constructed and the storage/release behaviors of Zn²⁺ ions are deeply investigated in these two typical structures. It should be mentioned that the 2D layered Na₅V₁₂O₃₂ and HNaV₆O₁₆·4H₂O with more effective path for Zn²⁺ diffusion exhibit higher ion diffusion coefficients than that of tunneled Na_0.76V₆O₁₅. As a result, Na₅V₁₂O₃₂ delivers higher capacity than that of Na_0.76V₆O₁₅, and a long‐term cyclic performance up to 2000 cycles at 4.0 A g^?1 in spite of its capacity fading. This work provides a new perspective of Zn²⁺ storage mechanism in aqueous ZIB systems.     

1H-NMR study of GM2 ganglioside: evidence that an interresidue amide-carboxyl hydrogen bond contributes to stabilization of a preferred conformation

Several properties of the exchangeable amide protons of the ganglioside G_M2 were studied in detail by¹H-NMR spectroscopy in fully deuterated dimethylsulfoxide [²H₆]DMSO/2% H₂O, and compared with data obtained for the simpler constituent glycosphingolipids G_A2 and G_M3. In addition to chemical shifts,³ J _2,HN coupling constants, and temperature shift coefficients, the kinetics of NH/²H chemical exchange were examined by following the disappearance of the amide resonances in [²H₆]DMSO/2%²H₂O. The results included observation of an increase in half-life of theN-acetylgalactosamine acetamido HN by more than an order of magnitude in G_M2 compared to G_A2, attributable to the presence of the additionalN-acetylneuraminic acid residue. Additional one-dimensional dipolar cross relaxation experiments were also performed on nonexchangeable protons of G_M2. The results of all of these experiments support a three-dimensional model for the terminal trisaccharide in which a hydrogen bond is formed between theN-acetylgalactosamine acetamido NH and theN-acetylneuraminic acid carboxyl group. The interaction is proposed to be of the -acceptor type, a possibility which has not yet been explored in the literature on carbohydrates. The proposed model is discussed in comparison with that of Sabesanet al. (1984,Can J Chem 62: 1034–45), and the models of G_M1 proposed more recently by Acquottiet al. (1990,J Am Chem Soc 112:7772–8) and Scarsdaleet al. (1990,Biochemistry 29:9843–55).     

Hydration of Escherichia coli lipids: Deuterium T1 relaxation time studies of phosphatidylglycerol,phosphatidylethanolamine and phosphatidylcholine

The hydration properties of Escherichia coli lipids (phosphatidylglycerol, phosphatidylethanolamine) and synthetic $\text{1,2-}\text{dioleoyl}\text{-sn-}\text{glycero-3-phosphocholine}$ in H₂O/²H₂O mixtures (9:1, v/v) were investigated with ²H-NMR. Comparison of the ²H₂O spin lattice relaxation time ($\text{T}{}_1$) as a function of the water content revealed a remarkable quantitative similarity of all three lipid-H₂O systems. Two distinct hydration regions could be discerned in the $\text{T}{}_1$ relaxation time profile. (1) A minimum of 11–16 water molecules was needed to form a primary hydration shell, characterized by an average relaxation time of $\text{T}{}_1\text{≈ 90}\text{ms}$. (2) Additional water was found to be in exchange with the primary hydration shell. The exchange process could be described in terms of a two-site exchange model, assuming rapid exchange between bulk water with $\text{T}{}_1\text{= 500}\text{ms}$ and hydration water with $\text{T}{}_1\text{= 80–120}\text{ms}$. Analysis of the linewidth and the residual quadrupole splitting (at low water content) confirmed the size of the primary hydration layer. However, each lipid-water system exhibited a somewhat different linewidth behavior, and a detailed molecular interpretation appeared to be preposterous.     

<Superscript>15</Superscript>N transverse relaxation measurements for the characterization of µs–ms dynamics are deteriorated by the deuterium isotope effect on <Superscript>15</Superscript>N resulting from solvent exchange

¹⁵N R₂ relaxation measurements are key for the elucidation of the dynamics of both folded and intrinsically disordered proteins (IDPs). Here we show, on the example of the intrinsically disordered protein α-synuclein and the folded domain PDZ2, that at physiological pH and near physiological temperatures amide—water exchange can severely skew Hahn-echo based ¹⁵N R₂ relaxation measurements as well as low frequency data points in CPMG relaxation dispersion experiments. The nature thereof is the solvent exchange with deuterium in the sample buffer, which modulates the ¹⁵N chemical shift tensor via the deuterium isotope effect, adding to the apparent relaxation decay which leads to systematic errors in the relaxation data. This results in an artificial increase of the measured apparent ¹⁵N R₂ rate constants—which should not be mistaken with protein inherent chemical exchange contributions, R_ex, to ¹⁵N R₂. For measurements of ¹⁵N R₂ rate constants of IDPs and folded proteins at physiological temperatures and pH, we recommend therefore the use of a very low D₂O molar fraction in the sample buffer, as low as 1%, or the use of an external D₂O reference along with a modified ¹⁵N R₂ Hahn-echo based experiment. This combination allows for the measurement of R_ex contributions to ¹⁵N R₂ originating from conformational exchange in a time window from µs to ms.     

Effect of pH on H-2H exchange,H2 production and H2 uptake,catalysed by the membrane-bound hydrogenase of Paracoccus denitrificans

(1) The kinetics of isotope exchange catalysed by the membrane-bound hydrogenase of Paracoccus denitrificans have been studied by measuring H²H, H₂ or ²H₂ produced when the enzyme catalyses the exchange between ²H₂ and H₂O or H₂ and ²H₂O. (2) In the ²H₂-H₂O system the measured rate of H₂ production was always higher than that of H²H. The $\text{H}{}_2\text{H}{}^2\text{H}$ ratio remained constant (about 1.70) in the protein concentration range 0.08–1.32 mg. The very rapid formation of H₂ with respect to H²H is consistent with the hypothesis of a heterolytic cleavage of ²H₂ into a deuteron and an enzyme hydride that can exchange with the solvent. (3) In the H₂-²H₂O system, the exchange rate was much lower than in the ²H₂-H₂O system, indicating a marked isotopic effect of ²H₂O. (4) The H-²H exchange activity, determined from the initial velocity of H²H formation, is optimal at pH 4.5. A second maximum of activity is observed at pH 8.3. The pH value of 4.5 is also the pH optimum for H₂ production while at pH 8.3–8.5 there is a maximum of H₂ oxidation activity. (5) In ordinary H₂O the K_m for hydrogen uptake estimated either from H₂ consumption or from benzyl viologen reduction was 0.06–0.07 μM for both H₂ and ²H₂ indicating a strong affinity of the enzyme for hydrogen at pH 8.3–8.5. Shifting from H₂O to ²H₂O does not affect the K_m of the enzyme for H₂ but lowers the V_max value about 10-fold. The K_m for benzyl viologen and methyl viologen was 0.08 and 2 mM, respectively.     

Synthesis, structure and solution chemistry of mixed-ligand oxovanadium(IV) and oxovanadium(V) complexes incorporating tridentate ONO donor hydrazone ligands

In the title family, the ONO donor ligands are the acetylhydrazones of salicylaldehyde (H₂L¹) and 2-hydroxyacetophenone (H₂L²) (general abbreviation, H₂L). The reaction of bis(acetylacetonato)oxovanadium(IV) with a mixture of tridentate H₂L and a bidentate NN donor [e.g., 2,2′-bipyridine(bpy) or 1,10-phenanthroline(phen), hereafter B] ligands in equimolar ratio afforded the tetravalent complexes of the type [V^IVO(L)(B)]; complexes (1)-(4) whereas, if B is replaced by 8-hydroxyquinoline(Hhq) (which is a bidentate ON donor ligand), the above reaction mixture yielded the pentavalent complexes of the type [V^VO(L)(hq)]; complexes (5) and (6). Aerial oxygen is most likely the oxidant (for the oxidation of V^IV → V^V) in the synthesis of pentavalent complexes (5) and (6). [V^IVO(L)(B)] complexes are one electron paramagnetic and display axial EPR spectra, while the [V^VO(L)(hq)] complexes are diamagnetic. The X-ray structure of [V^VO(L²)(hq)] (6) indicates that H₂L² ligand is bonded with the vanadium meridionally in a tridentate dinegative fashion through its phenolic-O, enolic-O and imine-N atoms. The general bond length order is: oxo < phenolato < enolato. The V-O (enolato) bond is longer than V-O (phenolato) bond by ∼0.07 Å and is identical with V-O (carboxylate) bond. ¹H NMR spectrum of (6) in CDCl₃ solution indicates that the binding nature in the solid state is also retained in solution. Complexes (1)-(4) display two ligand-field transitions in the visible region near 820 and 480 nm in DMF solution and exhibit irreversible oxidation peak near +0.60 V versus SCE in DMSO solution, while complexes (5) and (6) exhibit only LMCT band near 535 nm and display quasi-reversible one electron reduction peak near −0.10 V versus SCE in CH₂Cl₂ solution. The VO³⁺-VO²⁺E_1/2 values shift considerably to more negative values when neutral NN donor is replaced by anionic ON donor species and it also provides better VO³⁺ binding via phenolato oxygen. For a given bidentate ligand, E_1/2 increases in the order: (L²)²⁻ < (L¹)²⁻.     

NMR study of 17O from H217O in human erythrocytes

Human erythrocytes were incubated in a Ringer's solution enriched with 10–18% H₂¹⁷O. The longitudinal relaxation time (T₁) of the ¹⁷O was determined separately in samples of red cell suspesions, packed cells, and supernatant. The longitudinal relaxation of ¹⁷O in erythrocyte suspensions was non-exponential, reflecting water exchange across the cell membranes as well as relaxation processes inside and outside the cell.The T₁ of intracellular ¹⁷O is 4–5 times shorter than in the supernatant, similar to the enhancement of proton relaxation by hemoglobin in erythrocytes and free solution at the frequency applied (8.13 MHz). This datum is consistent with the thesis that hemoglobin modifies the NMR relaxation behavior of water inside cells and in free solution in the same way.The rate constant

for water exchange was calculated to be 60 and 107 s⁻¹ at 25 and at 37° C, respectively. The apparent activation energy for

over the temperature range 23–37° C was 8.7±1.0 kcal/mole.     

Direct assessment of water exchange on a Gd(III) chelate bound to a protein

 The Gd(III) complex of 4-pentylbicyclo[2.2.2]octane-1-carboxyl-di-l-aspartyl-lysine-derived DTPA, [GdL(H₂O)]^2–, binds to serum albumin in vivo, through hydrophobic interaction. A variable temperature ¹⁷O NMR, EPR, and Nuclear Magnetic Relaxation Dispersion (NMRD) study resulted in a water exchange rate of k ²⁹⁸ _ex=4.2×10⁶ s^–1, and let us conclude that the GdL complex is identical to [Gd(DTPA)(H₂O)]^2– in respect to water exchange and electronic relaxation. The effect of albumin binding on the water exchange rate has been directly evaluated by ¹⁷O NMR. Contrary to expectations, the water exchange rate on GdL does not decrease considerably when bound to bovine serum albumin (BSA); the lowest limit can be given as k _{ex, GdL-BSA}=k _ex, GdL / 2. In the knowledge of the water exchange rate for the BSA-bound GdL complex, the analysis of its NMRD profile at 35  °C yielded a rotational correlation time of 1.0 ns, one order of magnitude shorter than that of the whole protein. This value is supported by the longitudinal ¹⁷O relaxation rates. This indicates a remarkable internal flexibility, probably due to the relatively large distance between the protein- and metal-binding moieties of the ligand. Received: 25 June 1998 / Accepted: 11 August 1998     

Kinetics of Energetic O<Superscript>−</Superscript> Ions in the Discharge Plasmas of Water Vapor and H<Subscript>2</Subscript>O-Containing Mixtures

The process of relaxation of energetic O^– ions formed via dissociative attachment of electrons to molecules in the discharge plasmas of water vapor and H₂O: O₂ mixtures in a strong electric field is studied by the Monte Carlo method. The probability of energetic ions being involved in threshold ion–molecular processes is calculated. It is shown that several percent of energetic O^– ions formed via electron attachment to H₂O molecules in the course of plasma thermalization transform into OH^– ions via charge exchange or are destroyed with the formation of free electrons. The probabilities of charge exchange of O^– ions and electron detachment from them increase significantly (up to 90%) when O^– ions are formed via electron attachment to O₂ molecules in water vapor with an oxygen additive. This effect decreases with increasing oxygen fraction in the mixture but remains appreciable even when the fraction of H₂O molecules in the H₂O: O₂ mixture does not exceed several percent.     

Solution properties ofEscherichia coli-expressed VH domain of anti-neuraminidase antibody NC41

The V_H domain of anti-influenza neuraminidase antibody NC41, with and without a C-terminal hydrophilic marker peptide (FLAG^TM), has been expressed in high yield (15–27 mg/L) inEscherichia coli. Both forms were secreted into the periplasm where they formed insoluble aggregates which were solubilized quantitatively with 2 M guanidine hydrochloride and purified to homogeneity by ion-exchange chromatography. The V_H-FLAG was composed of three isoforms (pI values of 4.6, 4.9, and 5.3) and the V_H molecule was composed of two isoforms with pI values of 5.1 and 6.7; the difference between the V_H isoforms was shown to be due to cyclization of the N-terminal glutamine residue in the pI 5.1 isoform. At 20°C and concentrations of 5–10mg/ml the V_H domain dimerized in solution and then partly precipitated, resulting in the broadening of resonances in its¹H NMR spectrum. Reagents such as CHAPS,n-octylglucoside, and ethylene glycol, which presumably mask the exposed hydrophobic interface of the V_H molecule, prevented dimerization of the V_H and permitted good-quality NMR spectra on isotope-labeled protein to be obtained.     

Two dumbbell-like polyoxometalates constructed from capped molybdovanadate and transition metal complexes

Self assembly of NaVO₃, Na₂MoO₄·2H₂O and NiCl₂·6H₂O with the assistance of organic liginds under hydrothermal conditions results in two molybdovanadates [Ni(enMe)₂]₄{[Ni(enMe)₂(H₂O)]₂[Ni(enMe)₂][(V^VMo^VI₈V₄^IVO₄₀)(V^IVO)₂]₂}·10H₂O (1) and [Ni(enMe)₂]₅{[Ni(enMe)₂]₂[(V^VMo^VI₄Mo^V₄V₄^IVO₄₀)(V^IVO)₄]₂}·2H₂O (2), (enMe = 1,2-diaminopropane), which have been characterized by single crystal X-ray diffraction analysis, IR spectroscopy, and elemental analysis. Both of the two compounds exhibit dumbbell-like structures constructed from capped polyoxomolybdovanadates and [Ni(enMe)₂]²⁺ complexes. Polyoxoxanion 1 is composed of two bicapped Keggin-type anions [(V^VMo₈V₄^IVO₄₀)(V^IVO)₂]⁷⁻, one [Ni(enMe)₂]²⁺ bridging fragment and two decorated nickel(II) complexes. Polyxoxanion 2 consists of two tetracapped molybdenum-vanadium polyoxoanions [(V^VMo^VI₄Mo^V₄V₄^IVO₄₀)(V^IVO)₄]⁷⁻, one [Ni(enMe)₂]²⁺ bridging fragment and a nickel(II) decorated fragment. Polyxoxanions 1 and 2 are further linked to form three-dimensional supramolecular networks through extensive hydrogen bonding interactions. In addition, photocatalysis properties of these two compounds have been investigated.     

Effects of D2O on permeation and gating in the Ca2+-activated potassium channel fromChara

We studied the effects of H₂O/D₂O substitution on the permeation and gating of the large conductance Ca²⁺-activated K⁺ channels inChara gymnophylla droplet membrane using the patchclamp technique. The selectivity sequence of the channel was: K⁺>Rb⁺≫Li⁺, Na⁺, Cs⁺ and Cl⁻. The conductance of this channel in symmetric 100mm KCl was found to be 130 pS. The single channel conductance was decreased by 15% in D₂O as compared to H₂O. The blockade of channel conductance by cytosolic Ca²⁺ weakened in D₂O as a result of a decrease in zero voltage Ca²⁺ binding affinity by a factor of 1.4. Voltage-dependent channel gating was affected by D₂O primarily due to the change in Ca²⁺ binding to the channel during the activation step. The Hill coefficient for Ca²⁺ binding was 3 in D₂O and around 1 in H₂O. The values of the Ca²⁺ binding constant in the open channel conformation were 0.6 and 6 μm in H₂O and D₂O, respectively, while the binding in the closed conformation was much less affected by D₂O. The H₂O/D₂O substitution did not produce a significant change in the slope of channel voltage dependence but caused a shift as large as 60 mV with 1mm internal Ca²⁺.     

Polyoxometalate/laccase-mediated oxidative polymerization of catechol for textile dyeing

The synergistic effect between polyoxometalates (POMs), namely K₅[SiW₁₁V^VO₄₀]·11H₂O and H₅[PMo₁₀V^V ₂O₄₀]·13H₂O and laccase from ascomycete Myceliophthora thermophila has been employed for the first time in oxidative polymerization of catechol. Such a laccase-mediator system allowed the formation of a relatively high molecular weight polycatechol as confirmed by size exclusion chromatography and electrospray ionization mass spectrometry (ESI-MS) (3990 Da when using K₅[SiW₁₁V^VO₄₀]·11H₂O and 3600 Da with H₅[PMo₁₀V^V ₂O₄₀]·13H₂O). The synthesized polymers were applied as dyes for the dyeing of flax fabrics. The color intensity of flax fabrics colored with polymer solutions was evaluated by diffuse reflectance spectrophotometry via k/s measurements (+10% of fixation ratio). A new synthetic process allowed a dyeing polymer, provided upon flax coloration, better color fixation and color resistance when compared to that obtained by conventional synthesis with laccase solely or with addition of organic mediator (1-hydroxybenzotriazole).     

H2 oxidation,O2 uptake and CO2 fixation in hydrogen treated soils

In many legume nodules, the H₂ produced as a byproduct of N₂ fixation diffuses out of the nodule and is consumed by the soil. To study the fate of this H₂ in soil, a H₂ treatment system was developed that provided a 300 cm³ sample of a soil:silica sand (2:1) mixture with a H₂ exposure rate (147 nmol H₂ cm^–3hr^–1) similar to that calculated exist in soils located within 1–4 cm of nodules (30–254 nmol H₂ cm^–3hr^–1). After 3 weeks of H₂ pretreatment, the treated soils had a K_m and V_max for H₂ uptake (1028 ppm and 836 nmol cm^–3 hr^–1, respectively) much greater than that of control, air-treated soil (40.2 ppm and 4.35 nmol cm^–3 hr^–1, respectively). In the H₂ treated soils, O₂, CO₂ and H₂ exchange rates were measured simultaneously in the presence of various pH₂. With increasing pH₂, a 5-fold increase was observed in O₂ uptake, and CO₂ evolution declined such that net CO₂ fixation was observed in treatments of 680 ppm H₂ or more. At the H₂ exposure rate used to pretreat the soil, 60% of the electrons from H₂ were passed to O₂, and 40% were used to support CO₂ fixation. The effect of H₂ on the energy and C metabolism of soil may account for the well-known effect of legumes in promoting soil C deposition.     

Deuterium isotope effects on the central carbon metabolism of Escherichia coli cells grown on a D2O-containing minimal medium

In this paper it is demonstrated that cross-correlated time modulation of isotropic chemical shifts (`conformational exchange') leads to differential relaxation of double- and zero-quantum coherences, respectively. Quantitative information can be obtained from the time dependence of the interconversion between the two two-spin coherences 2I_xS_x and 2I_yS_y, induced by the differential relaxation. The effect is illustrated with an application to ¹³C,¹⁵N-labeled quail CRP2(LIM2), by studying ¹⁵N-¹H^N multiple-quantum relaxation. Significant cross-correlated fluctuations of isotropic chemical shifts were observed for residues which are part of a disordered loop region connecting two -strands in CRP2(LIM2). Differential ¹H^N and ¹⁵N exchange contributions to multiple-quantum relaxation observed at these sites illustrate the complex interplay between hydrogen bonding events and conformational reorientations in proteins.     

A new 3D transition-metal complex-templated framework based on Wells-Dawson polyoxometalate and copper(I)-pyrazine moieties

A new Wells-Dawson polyoxometalate-based compound, [Cu^II(2,2′-bipy)₂(H₂O)]₃[Cu^I₆(pz)₆(P₂W₁₈O₆₂)₂] (2,2′-bipy = 2,2′-bipyridine, pz = pyrizine) (1), has been hydrothermally synthesized and characterized by routine physical methods. Compound 1 exhibits a [Cu^II(2,2′-bipy)₂(H₂O)]²⁺ complex-templated 3D (3,4)-connected framework with the topology of (6³)(6²8⁴), which is built up by the cross-linking of porous P₂W₁₈-Cu layers and Cu^I-pz chains with Cu2 atoms as intersections. The transition-metal complex cation [Cu^II(2,2′-bipy)₂(H₂O)]²⁺ acts not only as a template locating in the voids of inorganic layers, but also as a charge-balance complex to make the 3D structure more steady. The electrochemistry property of compound 1 has also been discussed.