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1.
《Biochimica et Biophysica Acta (BBA)/General Subjects》1976,451(2):604-609
Shporer and Civan (Biochim. Biophys. Acta (1974) 354, 291–304) reported the effect of magnetic-field strength on the NMR relaxation times of 23Na in frog skeletal muscle. From these data, they estimated the correlation time τc for bound 23Na whose tumbling is severely restricted, and they suggested that the fraction of bound 23Na does not exceed some few percent of the total 23Na population. However, a step in their theoretical approach seems oversimplified. With an improved approach, we obtained an effective τc of 4–9 ns for bound 23Na. This value is some 10 times shorter than the corresponding value estimated by them from the same data. On the other hand, their conclusion concerning the amount of bound 23Na seems to remain valid. The origin of the observed difference between the two transverse relaxation times of tissue 23Na is also discussed. 相似文献
2.
Yoshihiro Hotta Hiroaki Ando Kazumi Takeya Jinsaku Sakakibara 《Molecular and cellular biochemistry》1994,139(1):59-70
The effects of the cardiac glycoside dihydroouabain (DHO), and the ericaceous toxin grayanotoxin-I (GTX-I) on myocardial cellular sodium (Nai) concentrations were investigated using sodium-23 nuclear magnetic resonance (23Na NMR) spectroscopy at 30°C in isolated perfused guinea-pig hearts. The Nai NMR signals from perfused Langendorff heart preparations were obtained by the modified inversion recovery (IR) method based on the previous observation that the spin-lattice relaxation time (T1) of the Nai (25 or 34 msec at 8.46 Tesla (T)) is much faster than that of extracellular sodium (64 msec at 9.4 T). Nai was estimated from the calibration curve of the frequency area of the23Na NMR FT spectra plotted against the standard Na concentration. The Nai concentration of the heart increased concomitantly with the positive inotropic effects (PIE) of DHO, GTX-I and monensin (MON). The cumulative sequential addition of DHO (5×10–6 M), GTX-I (7×10–8 M) and MON (5×10–6 M), each of which alone induced no appreciable PIE, produced a 22% elevation in Nai concentration relative to that of the control (100%) accompanying a PIE of 44%. The mechanism of this Nai elevation induced by combinational addition of DHO, GTX-I and MON may be mediated as follows: GTX-I increases the net Na-influxvia Na+ channels; DHO inhibits the pumping out of Na+ from the cell; and MON transports external Na+ into the cell, acting as a sodium ionophore. Consequently, these drugs act synergistically to increase the Nai, thereby increasing the intracellular Ca2+ concentrationvia Na+–Ca2+ exchange. 相似文献
3.
Mordechai Shporer Mortimer M. Civan 《Biochimica et Biophysica Acta (BBA)/General Subjects》1974,354(2)
Pulsed NMR techniques have been applied to the study of the relaxation parameters characterizing 23Na within frog striated muscle. Experiments were performed at 3°C, 22–24°C and 39°C at a Larmor frequency of 15.7 MHz; at 22–24°C, measurements were obtained both at 15.7 MHz and at 7.85 MHz.As previously reported, only a single spine-lattice relaxation time (T1) was observed, but both slow (T2)I and fast (T2)II components of the spin-spin relaxation time were measured. The effect of temperature (θ) upon (1/T1) was qualitatively similar to that reported for 23Na in free solution; (θ) did not significantly affect (1/T2) over the range of temperatures studied. (1/T2)I, and to a lesser degreee, (1/T1) exhibited a modest inverse dependence of doubtful significance on the Larmor frequency.The data are examined within the framework of a simple specific model; a conservative values in assumed for the quadrupolar coupling constant characterizing immobilized intracellular Na+. Within this framework, the results suggest that the fraction of bound ions whose molecular tumbling is severely restricted does not exceed some few percent of the total sodium population. 相似文献
4.
Summary The three-compartment model (paper I) described turgor pressure relaxations as a sum of two exponential functions. The predicted shape of the curves could be confirmed inChara corallina by improving the recording and processing of data. An evaluation on the basis of the three-compartment model provided values for the hydraulic conductivity of the plasmalemma ofLp
p=2×10–5 to 4×10–5 cm sec–1 bar–1 and ofLp
i=3×10–5 to 1×10–4 cm sec–1 bar–1 for the tonoplast (assuming the area to be 90% of the plasmalemma area). The mean proportion of the total volume occupied by the cytoplasm was calculated to be 9%. This value is consistent with the concept of a highly vacuolated cell. Other explanations for the biphasic relaxation curves are discussed. 相似文献
5.
Soluble cytochrome c-554 (M
r 10 kDa) is purified from the green sulfur bacterium Chlorobium tepidum. Its midpoint redox potential is determined to be +148 mV from redox titration at pH 7.0. The kinetics of cytochrome c-554 oxidation by a purified reaction center complex from the same organism were studied by flash absorption spectroscopy at room temperature, and the results indicate that the reaction partner of cytochrome c-554 is cytochrome c-551 bound to the reaction center rather than the primary donor P840. The second-order rate constant for the electron donation from cytochrome c-554 to cytochrome c-551 was estimated to be 1.7×107 M–1 s–1. The reaction rate was not significantly influenced by the ionic strength of the reaction medium.This revised version was published online in October 2005 with corrections to the Cover Date. 相似文献
6.
Summary Basal lateral membrane vesicles were isolated from rat intestinal epithelial cells. The sodium potassium triphosphatase (Na/K-ATPase) of these plasma membranes has been characterized by (1) the molecular weight of the phosphorylated intermediate, (2) the sensitivity of the phosphorylated intermediate to hydroxylamine, (3) its ouabain binding constants, and (4) its susceptibility to digestion by pronase. The phosphorylated intermediate was shown by SDS polyacrylamide gel electrophoresis to be a protein of 100,000 Daltons apparent mol wt. Its extensive hydrolysis in hydroxylamine demonstrated that it was an acyl phosphate. The isolated basal lateral membranes bound ouabain with a dissociation constant,K
m
(1.5×10–5
m), similar to the inhibitory constantK
I
(3×10–5
m), measured for ouabain inhibition of the Na/K-ATPase activity. The association rate constant measured for ouabain binding at 22°C was 1.3×103
m
–1 sec–1 and is similar to the association rate constants reported for other tissues and species. The high dissociation rate constant, 3.6×10–2 sec–1, is consistent with the insensitivity of the rat to ouabain. Digestion of the intact cells by pronase yielded basal lateral membranes in which the Na/K-ATPase had been unaffected. The phosphorylated intermediate ran as a sharp band at 100,000 Daltons on electrophoresis, and the ouabain dissociation constant appeared to be unchanged. In these membranes, protein stains of polyacrylamide gels revealed digestion of the major high mol wt proteins including the major protein at 100,000 Daltons. This suggests that the Na/K-ATPase represent a minor component, less than 1%, of the basal lateral membrane protein. From these characteristics of the phosphorylated intermediate and the ouabain binding constants, we conclude that the Na/K-ATPase of the basal lateral membranes of rat intestinal epithelial cells is similar to that found in other tissues and species. Estimates of the number of pump sites and the turnover number predict rates of Na transport that are consistent with observed values.This paper is dedicated to the memory of Professor David H. Smyth, FRS, who died on September 10, 1979. 相似文献
7.
Summary The charge-pulse technique has been used previously for the study of quasistationary processes in membranes which required only a moderate time resolution. It is shown here that a time resolution of about 400 nsec may be achieved with this technique and that it may be applied to the kinetic analysis of carrier-mediated ion transport. By this method we have studied the transport of alkali ions through optically black monoolein membranes in the presence of the ion carrier valinomycin. All three relaxation processes that are predicted by theory have been resolved. From the relaxation times and the relaxation amplitudes the rate constants for the association (k
R
) and the dissociation (k
D
) of the ioncarrier complex, as well as the translocation rate constants of the complex (k
MS
) and the free carrier (k
S
) could be obtained. For 1m Rb+ at 25° C the values arek
R
=3×105
m
–1 sec–1,k
D
=2×105 sec–1,k
MS
=3×105 sec–1,k
S
=4×104 sec–1. The activation energies of the single rate constants which have been estimated from experiments at two different temperatures range between 50 and 90 kJ/mol. 相似文献
8.
Summary A new control policy for the on-line optimization of the nutrient supply in bakers yeast process is proposed. A feed rate corresponding to minimal substrate uptake time was shown to be optimal for cell yield and specific growth rate. Cultivation results of baker's yeast are presented.Nomenclature c
glucose concentration in wort (mol.l–1)
- C
total glucose used (mol)
- ce
ethanol concentration in wort (mg.l–1)
- cp
glucose concentration in fresh medium (mol.l–1)
- dt/dc
glucose consumption time (sec.mol–1)
- F
substrate feed rate (litre.hr–1)
- qc
glucose uptake rate (mol.hr–1)
- Qc
specific glucose uptake rate (moll.g–1.hr–1)
- qO2
oxygen uptake rate (mol.hr–1)
- QO2
specific oxygen uptake rate (mol.g–1.hr–1)
- rx
productivity (g.l–1.hr–1)
- t
time (hr)
- x
biomass concentration (g.l–1)
- X
total biomass (g)
- Yx/c
cell yield (g.g–1): (g.mol–1)
- Yo/c
consumed oxygen to glucose ratio (mol.mol–1) 相似文献
9.
Alexander V. Itkes Kyril T. Turpaev Olga N. Kartasheva Constantin A. Kafiam Evgenii S. Severin 《Molecular and cellular biochemistry》1984,59(1-2):165-171
Summary Dihydrofolate synthetase (EC 6.3.2.12) from N. gonorrhoeae was isolated and enzyme characteristics were determined. The purified enzyme was found quite stable when stored at –60 °C. About 50% of the enzyme activity wag destroyed within 6 weeks when kept at 4 °C. Maximum velocity was observed at pH 9.3. The enzyme required a monovalent cation, K+ or NH4
+ , and divalent cation, Mg2+ or Mn2+ for its function. ATP at 5 mM concentration gave maximum activity. Km values for dihydropteroate and L-glutamate at pH 9.3 were 3.5 × 10–5 M and 6.5 × 10–4 M, respectively. Patterns of product inhibition by dihydrofolate were found to be non-competitive with respect to dihydropteroate, having a Ki value of 5.1 ± 0.8 × 10–4 M, and competitive with respect to L-glutamate, having a Ki value of 6.2 × 10–4 M. 相似文献
10.
Yoshinobu Tsuchiya Naomichi Nishio Shiro Nagai 《Applied microbiology and biotechnology》1980,9(2):121-127
Summary In the productions of biomass and vitamin B12 using methanol as the sole carbon source, it is necessary to use a medium in which methanol is the growth limiting substrate. Other inorganic salts should be in slight excess so that the yield of cells and the intracellular content of vitamin B12 do not vary. From basic principles of chemostat culture, a medium was optimized for Pseudomonas AM-1 a methanol utilizing bacterium, for the concentrations of various inorganic salts. This was done in a series of chemostat cultures at a dilution rate of 0.1 h–1. Optimum amounts of NH4
+, PO4
3- and Mg2+ were estimated from the minimum concentration of the salt at which methanol became growth limiting. The optimum concentrations of Ca2+, Fe2+, Mn2+, and Zn2+ as a group were determined in the same way. Cu2+, Mo6+, Co2+ and B3+ are required at concentrations of g/l and they were not studied as these very low level can be introduced as contaminants from other salts. The optimum medium composition (in g/l) was as follows: (NH4)2SO4, 1.0; H3PO4, 75×10–3; MgSO4 · 7H2O, 30×10–3; CaCl2 · 2H2O, 3.3×10–3; FeSO4 · 7H2O, 1.3×10–3, MnSO4 · 4H2O, 0.13×10–3; ZnSO4 · 7H2O, 0.13×10–3; CuSO4 · 5H2O, 40×10–6; Na2MoO4, 40×10–6; CoCl2 · 6H2O, 40×10–6; H3BO3, 30×10–6 and methanol 4. 相似文献
11.
Ahlert Schmidt 《Archives of microbiology》1973,93(1):29-52
Summary During sulfate reduction in a cell-free system ofChlorella activated sulfate of APS is transferred to a thiosulfonate reductase. The sulfate thus bound to the thiosulfonate reductase (i.e. bound sulfite) is reduced to bound sulfide in a ferredoxin dependent reaction. This bound sulfide can be used with O-acetylserine as acceptor for cysteine biosynthesis; serine and O-phosphoserine are not used. An assaysystem for thiosulfonate reductase activity using methylviologen dependent reduction of S2O4
2– to S2– is developed and a procedure for isolating thiosulfonate reductase fromChlorella cells is presented. During isolation of thiosulfonate reductase a low weight molecular factor, needed for optimal enzyme activity was lost. The bound sulfite seems to be attached to this factor. Reduction of APS or GS-SO3H to the level of S2– is inhibited by cysteine. 50% inhibition of GS-SO3H reduction was found at a molar cysteine concentration of 6.8×10–5.Abbreviations APS
adenosine-phosphosulfate
- PAPS
3-phosphoadenosine-5-phosphosulfate
- GSH
reduced glutathion
- GS-SO3H
S-sulfoglutathion
- fd
ferredoxin
- Mv
methylviologen
- DTT
dithiothreitol 相似文献
12.
In this work, we derive an analytical expression for the relaxation time τ as a function of temperature T for myoglobin protein (Mb, PDB:1MBN) in the high temperature limit (T > Tg = 200 K). The method is based on a modified version of the Adam–Gibbs theory (AG theory) for the glass transition in supercooled liquids and an implementation of differential geometry techniques. This modified version of the AG theory takes into account that the entropic component in protein's denaturation has two major sources: a configurational contribution ΔSc due to the unfolding of the highly ordered native state N and a hydration contribution ΔShyd arising from the exposure of non-polar residues to direct contact with solvent polar molecules. Our results show that the configurational contribution ΔSc is temperature-independent and one order of magnitude smaller than its hydration counterpart ΔShyd in the temperature range considered. The profile obtained for log τ(T) from T = 200 K to T = 300 K exhibits a non-Arrhenius behavior characteristic of α relaxation mechanisms in hydrated proteins and glassy systems. This result is in agreement with recent dielectric spectroscopy data obtained for hydrated myoglobin, where at least two fast relaxation processes in the high temperature limit have been observed. The connection between the relaxation process calculated here and the experimental results is outlined. 相似文献
13.
Zand Robert Marcelo Cynthia L. MacKenzie Robert Georgic Lynn Maclean Derrick Dunham William R. 《Neurochemical research》1997,22(4):437-443
The retention rate of the spin label 3-isothiocyanto methyl-2,2,5,5-tetramethyl-1-pyrrolidinyl oxyl spin label (proxyl) attached to the porcine N-acetyl-NPY peptide and the porcine N-acetyl-D-Trp32-NPY peptide at Lys4 was investigated using SK-N-MC neuroblastoma cell membranes containing the Y1 receptor. The release rate of the spin labeled peptides was monitored by electron spin resonance and the KD was determined by a direct radiolabeled NPY displacement binding assay. The analyses show that for the porcine [Ac-Tyr1N4-proxyl]-NPY, the KD was 8 × 10–10 M and koff was 2.7 × 10–4 sec–1 yielding a value for kon of 3.3 × 105 sec–1 M–1. The [Ac-Tyr1, N4-proxyl,-D-Trp32]-NPY antagonist ligand had a value of KD equal to 1.35 × 10–7 M and koff was 1.7 × 10–4 sec–1 leading to a value for kon of 1.2 × 103 sec–1 M–1. The difference in the kon rates of two orders of magnitude is interpreted as demonstrating the N-acetyl-N4 proxyl-D-Trp32-NPY ligand binding transition state to be of higher energy then for the unmodified NPY amino acid sequence. 相似文献
14.
The turnover times of glucose, averaged for 0–10 m in the upper waters of Lake Kinneret and measured by the addition of single or multiple concentrations of substrate, ranged from 23 to 188 hours and 1 to 87 hours respectively. Potential uptake rates (estimated as Vmax) ranged from 0.095 to 1.94 µg glucose l–1h–1, while measured uptake rates varied from 0.09 to 1.1 µg glucose l–1h–1. Concentrations of dissolved carbohydrates and glucose averaged 0.71 mg glucose equivalents l–1 and 39 µg glucose l–1 respectively. No evident relationships between glucose cycling and any fractions of dissolved organic matter, phytoplankton biomass or primary productivity were found. Turnover times were generally most rapid immediately after the decline of the spring Peridinium bloom. The respiration percentage of incorporated glucose ranged from 25% to 61% with highest values during the summer months. Respiration may be influenced by the nature of the indigenous bacterial population as well as by temperature. Daily heterotrophic glucose carbon uptake was about 9% of the photosynthetic incorporation and could provide a bacterial yield of about 7 × 104 ml–1d–1. 相似文献
15.
J. F. García-Díaz G. Klemperer L. M. Baxendale A. Essig 《The Journal of membrane biology》1986,92(1):37-46
Summary Cell Na activity,a
Na
c
, was measured in the short-circuited frog skin by simulaneous cell punctures from the apical surface with open-tip and Na-selective microelectrodes. Skins were bathed on the serosal surface with NaCl Ringer and, to reduce paracellular conductance, with NaNO3 Ringer on the apical surface. Under control conditionsa
Na
c
averaged 8±2mm (n=9,sd). Apical addition of amiloride (20 m) or Na replacement reduceda
Na
c
to 3mm in 6–15 min. Sequential decreases in apical [Na] induced parallel reductions ina
Na
c
and cell current,I
c
. On restoring Na after several minutes of exposure to apical Na-free solutionI
c
rose rapidly
to a stable value whilea
Na
c
increased exponentially, with a time constant of 1.8±0.7 min (n=8). Analysis of the time course ofa
Na
c
indicates that the pump Na flux is linearly related toa
Na
c
in the range 2–12mm. These results indicate thata
Na
c
plays an important role in relating apical Na entry to basolateral active Na flux. 相似文献
16.
Sulfate reduction and S-oxidation in a moorland pool sediment 总被引:3,自引:2,他引:1
In an oligotrophic moorland pool in The Netherlands, S cycling near the sediment/water boundary was investigated by measuring (1) SO4
2– reduction rates in the sediment, (2) depletion of SO4
2– in the overlying water column and (3) release of35S from the sediment into the water column. Two locations differing in sediment type (highly organic and sandy) were compared, with respect to reduction rates and depletion of SO4
2– in the overlying water.Sulfate reduction rates in sediments of an oligotrophic moorland pool were estimated by diagenetic modelling and whole core35SO4
2– injection. Rates of SO4
2– consumption in the overlying water were estimated by changes in SO4
2– concentration over time in in situ enclosures. Reduction rates ranged from 0.27–11.2 mmol m–2 d–1. Rates of SO4
2– uptake from the enclosed water column varied from –0.5, –0.3 mmol m–2 d–1 (November) to 0.43–1.81 mmol m–2 d–1 (July, August and April). Maximum rates of oxidation to SO4
2– in July 1990 estimated by combination of SO4
2– reduction rates and rates of in situ SO4
2– uptake in the enclosed water column were 10.3 and 10.5 mmol m–2 d–1 at an organic rich and at a sandy site respectively.Experiments with35S2– and35SO4
2– tracer suggested (1) a rapid formation of organically bound S from dissimilatory reduced SO4
2– and (2) the presence of mainly non SO4
2–-S derived from reduced S transported from the sediment into the overlying water. A35S2– tracer experiment showed that about 7% of35S2– injected at 1 cm depth in a sediment core was recovered in the overlying water column.Sulfate reduction rates in sediments with higher volumetric mass fraction of organic matter did not significantly differ from those in sediments with a lower mass fraction of organic matter.Corresponding author 相似文献
17.
Kai-Wen Huang Hsin-Hsien Chen Hong-Chang Yang Herng-Er Horng Shu-Hsien Liao Shieh Yueh Yang Jen-Jie Chieh Li-Ming Wang 《PloS one》2012,7(10)
In this study, we report the spin-lattice relaxation rate of hepatocellular carcinoma (HCC) and normal liver tissue in rats using a high-Tc superconducting quantum interference device (SQUID) based nuclear magnetic resonance (NMR) spectrometer. The resonance spectrometer used for discriminating liver tumors in rats via the difference in longitudinal relaxation time in low magnetic fields was set up in a compact and portable magnetic shielding box. The frequency-domain NMR signals of HCC tissues and normal liver tissues were analyzed to study their respective longitudinal relaxation rate T1
−1. The T1
−1 of liver tissues for ten normal rats and ten cancerous rats were investigated respectively. The averaged T1
−1 value of normal liver tissue was (6.41±0.66) s−1, and the averaged T1
−1 value of cancerous tissue was (3.38±0.15) s−1. The ratio of T1
−1 for normal liver tissues and cancerous liver tissues of the rats investigated is estimated to be 1.9. Since this significant statistical difference, the T1
−1 value can be used to distinguish the HCC tissues from normal liver tissues. This method of examining liver and tumor tissues has the advantages of being convenient, easy to operate, and stable. 相似文献
18.
Summary The changes in Na current during development were studied in the dorsal root ganglion (DRG) cells using the whole-cell patch-clamp technique. Cells obtained from rats 1–3 and 5–8 days after birth were cultured and their Na currents were compared. On top of the two types of Na currents reported in these cells (fast-FA current and slow-S current) a new fast current was found (FN). The main characteristics of the three currents are: (i) The voltages of activation are –37, –36, and –23 mV for the FN, FA and S currents, respectively. (ii) The activation and inactivation kinetics of FN and FA currents are about five times faster than those of the S current. (iii) The voltages at which inactivation reaches 50% are –139, –75 and –23 mV for the FN, FA and S currents, respectively.The kinetics and voltage-dependent parameters of the three currents and their density do not change during the first eight days after birth. However, their relative frequency in the cells changes. In the 1–3 day-old rats the precent of cells with S, FA, and mixed S+FN currents is 22, 18, and 60% of the cells, respectively. In the 5–8 day-old, the percent of cells with S, FA, and FN+S is 10, 66 and 22%. The relative increase in the frequency of cells with FA current during development can contribute to the ease of action potential generation compared with cells with FN currents, which are almost completely inactivated under physiological conditions. The predominance of FA cells also results in a significant decrease in the relative frequency of cells with the high-threshold, slow current.Antibodies directed against a part of the S4 region of internal repeat I of the sodium channel (C
1
+
, amino acids 210–223, eel channel numbering) were found to shift the voltage dependence of FA current inactivation (but not of FN or S currents) to more negative potentials. The effect was found only when the antibodies were applied externally. The results suggest that FN, FA and S types of Na currents are generated by channels, which are different in the topography of the C
1
+
region in the membrane. 相似文献
19.
Yusuke Miyashita Tetsuichi Wazawa George Mogami Satoshi Takahashi Yoshihiro Sambongi Makoto Suzuki 《Biophysical journal》2013,104(1):163-172
We investigate the hydration state of horse-heart cytochrome c (hh cyt c) in the unfolding process induced by trifluoroacetic acid (TFA). The conformation of hh cyt c changes from the native (N) state (2.9 < pH < 6.0) to the acid-unfolded (UA) state (1.7 < pH < 2.0) to the acid-induced molten globule (A) state (pH ∼1.2). Hydration properties of hh cyt c during this process are measured at 20°C by high-resolution dielectric relaxation (DR) spectroscopy, UV-vis absorbance, and circular dichroism spectroscopy. Constrained water of hh cyt c is observed at every pH as an ∼5-GHz Debye component (DC) (DR time, τD ∼30 ps) and its DR amplitude (DRA) is increased by 77% upon N-to-UA transition, when pH changes from 6.0 to 2.0. Even in the N state, the DRA of the constrained-water component is found to be increased by 22% with decreasing pH from 6.0 to 2.9, suggesting an increase in the accessible surface area of native hh cyt c. Moreover, hypermobile water around native hh cyt c is detected at pH 6.0 as a 19-GHz DC (τD ∼ 8.4 ps < τDW = 9.4 ps), but is not found at other pH values. The DRA signal of constrained water is found to return to the pH 2.9 (N-state) level upon UA-to-A transition. Fast-response water (slightly slower than bulk) around A-state hh cyt c is detected at pH 1.2, and this suggests some accumulation of TFA− ions around the peptide chain. Thus, this high-resolution DR spectroscopy study reveals that hh cyt c exhibits significant hydration-state change in the TFA-unfolding process. 相似文献
20.
The physiological ligands for Na,K-ATPase (the Na,K-pump) are ions, and electrostatic forces, that could be revealed by their ionic strength dependence, are therefore expected to be important for their reaction with the enzyme. We found that the affinities for ADP3−, eosin2−, p-nitrophenylphosphate, and Vmax for Na,K-ATPase and K+-activated p-nitrophenylphosphatase activity, were all decreased by increasing salt concentration and by specific anions. Equilibrium binding of ADP was measured at 0–0.5 M of NaCl, Na2SO4, and NaNO3 and in 0.1 M Na-acetate, NaSCN, and NaClO4. The apparent affinity for ADP decreased up to 30 times. At equal ionic strength, I, the ranking of the salt effect was NaCl ≈ Na2SO4 ≈ Na-acetate < NaNO3 < NaSCN < NaClO4. We treated the influence of NaCl and Na2SO4 on K
diss for E·ADP as a “pure” ionic strength effect. It is quantitatively simulated by a model where the binding site and ADP are point charges, and where their activity coefficients are related to I by the limiting law of Debye and Hückel. The estimated net charge at the binding site of the enzyme was about +1. Eosin binding followed the same model. The NO3
− effect was compatible with competitive binding of NO3
− and ADP in addition to the general I-effect. K
diss for E·NO3 was ∼32 mM. Analysis of Vmax/K
m for Na,K-ATPase and K+-p-nitrophenylphosphatase activity shows that electrostatic forces are important for the binding of p-nitrophenylphosphate but not for the catalytic effect of ATP on the low affinity site. The net charge at the p-nitrophenylphosphate-binding site was also about +1. The results reported here indicate that the reversible interactions between ions and Na,K-ATPase can be grouped according to either simple Debye-Hückel behavior or to specific anion or cation interactions with the enzyme. 相似文献