Essential sulfhydryl group of malic enzyme fromEscherichia coli

The activity of malic enzyme fromEscherichia coli was unaffected by the monovalent cations Na⁺ or Li⁺ at 10 mM. At 100 mM, Li⁺ or Na⁺ inhibited the enzyme activity by 88% and 83%, respectively. However, the enzyme activity was stimulated by 40–80-fold with 10 mM K⁺, Rb⁺, Cs⁺, or NH ₄ ⁺ . Less stimulation was observed with 100 mM of these stimulating cations. The stimulatory effect was lost after the enzyme was dialyzed against Tris-Cl buffer, but was regained after incubating the dialyzed enzyme with dithiothreitol. The regenerated enzyme was inactivated by 5,5′-dithiobis(2-nitrobenzoic acid). The resulting inactive thionitrobenzoyl enzyme could be regenerated to the active thiol-enzyme by eithiothreitol or converted to the inactive thiocyanoylated enzyme by KCN. The thiocyanoylated enzyme was insensitive to K⁺ stimulation, which suggested the essentiality of the sulfhydryl groups of theE. coli malic enzyme.     

Induction of Myxospore Formation in Stigmatella aurantiaca (Myxobacterales) by Monovalent Cations

Suspension cultures of Stigmatella aurantiaca can be induced to form myxospores by addition of the monovalent cations Li⁺, Na⁺, NH₄⁺, K⁺, or Rb⁺.     

Plasmalemmal voltage-activated K + currents in protoplasts from tobacco BY-2 cells: possible regulation by actin microfilaments?

Summary.  Plasmalemmal ionic currents from enzymatically isolated protoplasts of suspension-cultured tobacco ‘Bright Yellow-2’ cells were investigated by whole-cell patch-clamp techniques. In all protoplasts, delayed rectifier outward K⁺ currents having sigmoidal activation kinetics, no inactivation, and very slow deactivation kinetics were activated by step depolarization. Tail current reversal potentials were close to equilibrium potential E_K when external [K⁺] was either 6 or 60 mM. Several channel blockers, including external Ba²⁺, niflumic acid, and 5-nitro-2-(3-phenylpropylamino)-benzoic acid, inhibited this outward K⁺ current. Among the monovalent cations tested (NH₄ ⁺, Rb⁺, Li⁺, Na⁺), only Rb⁺ had appreciable permeation (P_Rb/P_K ⁼ 0.7). In addition, in 60 mM K⁺ solutions, a hyperpolarization-activated, time-dependent, inwardly rectifying K⁺ current was observed in most protoplasts. This inward current activated very slowly, did not inactivate, and deactivated quickly upon repolarization. The tail current reversal potential was very close to E_K, and other monovalent cations (NH₄ ⁺, Rb⁺, Li⁺, Na⁺) were not permeant. The inward current was blocked by external Ba²⁺ and niflumic acid. External Cs⁺ reversibly blocked the inward current without affecting the outward current. The amplitude of the inward rectifier K⁺ current was generally small compared to the amplitude of the outward K⁺ current in the same cell, although this was highly variable. Similar amplitudes for both currents occurred in only 4% of the protoplasts in control conditions. Microfilament-depolymerizing drugs shifted this proportion to about 12%, suggesting that microfilaments participate in the regulation of K⁺ currents in tobacco ‘Bright Yellow-2’ cells. Received December 7, 2001; accepted April 15, 2002; published online July 4, 2002 RID="*" ID="*" Correspondence and reprints: Pharmacologie et Physicochimie, UMR CNRS 7034, Faculté de Pharmacie, Université Louis Pasteur, 74 route du Rhin, BP 24, 67401 Illkirch, France. Abbreviations: TBY-2 Tobacco ‘Bright Yellow-2’; DHCB dihydrocytochalasin B; I_Kin inward rectifier K⁺ current; I_Kout outward K⁺ current; MFs microfilaments; MTs microtubules; NPPB 5-nitro-2-(3-phenylpropylamino)-benzoic acid.     

Stimulating effects of monovalent cations on activator-dissociated and activator-associated states of Ca2+-ATPase in human erythrocytes

The additional activation by monovalent cations of the $\text{(Ca}{}^{\mathrm{2+}}\text{+ Mg}{}^{\mathrm{2+}}\text{)-dependent}$ ATPase (ATP phosphohydrolase, EC 3.6.1.3) in human erythrocyte membranes was studied.The Ca²⁺-ATPase occurs in two different states. In the A-state the enzyme is virtually free of protein activator and the kinetics of Ca²⁺ activation is characterized by low apparent Ca²⁺ affinity and low maximum activity. In the B-state the enzyme is associated with activator and the kinetics is characterized by high Ca²⁺ affinity and high maximum activity.At optimum concentrations of Ca²⁺ the additional activation of the B-state by K⁺, NH₄⁺, Na⁺ and Rb⁺ exceeded the corresponding activations of the A-state, and half-maximum activations by K⁺, NH₄⁺, and Na⁺ were achieved at lower concentrations in the B-state than in the A-state. Li⁺ and Cs⁺ activated the two states almost equally but maximum activation was obtained at lower cation concentrations in the B-state than in the A-state.The activation of the B-state by the various cations decreased in the order $\text{K}{}^{\mathrm{+}}\text{> NH}{}_4{}^{\mathrm{+}}\text{> Na}{}^{\mathrm{+}}\text{= Rb}{}^{\mathrm{+}}\text{> Li}{}^{\mathrm{+}}\text{= Cs}{}^{\mathrm{+}}$. The A-state was activated almost equally by K⁺, Na⁺, NH₄⁺, and Rb⁺ and to a smaller extent by Li⁺ and Cs⁺.At sub-optimum concentrations of Ca²⁺ high concentrations of monovalent cations (100 mM) activated the Ca²⁺-ATPase equally in the A-state and the B-state. In the absence of Ca²⁺ the monovalent cations inhibited the Mg²⁺-dependent ATPase in both types of membranes. This dependence on Ca²⁺ indicates that the monovalent cations interact with the Ca²⁺ sites in the B-state.The results suggest that K⁺ or Na⁺, or both, contribute to the regulation of the Ca²⁺ pump in erythrocytes.     

Influence of Certain Cations on Activity of Succinyl CoA Synthetase From Tobacco

Succinyl CoA synthetase from Nicotiana tabacum exhibited a requirement for univalent and divalent cations. Mn²⁺ replaced Mg²⁺ in the assay medium and Co²⁺ and Ca²⁺ partially replaced Mg²⁺. Addition of Zn²⁺ resulted in no enzyme activity. The enzyme was activated by univalent cations K⁺, Rb⁺, NH₄⁺, and Na⁺; Li⁺ showed little or no activation. Maximum enzyme activity varied significantly with potassium salts of different anions. Greatest activation was obtained with K₃PO₄ and, respectively, KCl, KNO₃, K₂SO₄ and KF exhibited steadily decreasing enzyme activation.     

Effect of NH+4 and K+ on the activity of the ribosomal subunits in the EF-G- and EF-T-dependent GTR hydrolysis

$\text{E. coli}$ 50S ribosomal subunits show in the absence of 30S subunits and at low NH₄⁺ or K⁺ high turnover activity in EF-G-dependent GTP hydrolysis which is inhibited by increasing concentrations of monovalent cations. At 80 mM NH₄⁺ or K⁺ this activity is already 70–80% inhibited. This effect is reversed by 30S which are stimulatory with an optimum at about 80 mM for NH₄⁺ and 20–40 mM for K⁺. At low NH₄⁺ or K⁺ (<5 mM) stimulation by 30S of maximal 50S activity depends on the [EF-G]/[50S]. Unlike EF-G, EF-T does not show any Phe-tRNA-dependent GTPase activity with 50S alone even at low concentrations of NH₄⁺ or K⁺.     

Physical and kinetic properties of sheep liver pyridoxine kinase

Pyridoxine kinase purified from sheep liver was found to consist of a single polypeptide chain with a molecular weight of 60,000 as determined by gel filtration, sedimentation equilibrium ultracentrifugation, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The isoelectric pH of the enzyme was 5.1, and the pH optimum was between 5.5 and 6.0. The enzyme required divalent cations for activity. At cation concentrations of 80 μm, the enzyme activity with each cation was in the order of Zn²⁺ > Mn²⁺ > Mg²⁺. At cation concentrations of 400 μm, the enzyme activity with each cation was in the order of Mn²⁺ > Zn²⁺ > Mg²⁺. Excess free divalent cation inhibited the enzyme. Pyridoxine kinase also required monovalent cations. The enzyme activation was greatest with K⁺, then Rb⁺ and NH₄⁺, whereas the enzyme had very little activity with Na⁺, Li⁺, or Cs⁺. Na⁺ did not interfere with the activation by K⁺. The activation of the kinase by K⁺, NH₄⁺, and Rb⁺ followed Michaelis-Menten kinetics, and the apparent K_m values for the cations were 8.9, 3.7, and 5.3 mm, respectively. Increasing the potassium concentration lowered the apparent K_m value of the enzyme for pyridoxine and had little or no effect on the K_m for ZnATP^2? or the V of the kinase-catalyzed reaction.     

Essential sulfhydryl group of malic enzyme fromEscherichia coli

The activity of malic enzyme fromEscherichia coli was unaffected by the monovalent cations Na⁺ or Li⁺ at 10 mM. At 100 mM, Li⁺ or Na⁺ inhibited the enzyme activity by 88% and 83%, respectively. However, the enzyme activity was stimulated by 40–80-fold with 10 mM K⁺, Rb⁺, Cs⁺, or NH ₄ ⁺ . Less stimulation was observed with 100 mM of these stimulating cations. The stimulatory effect was lost after the enzyme was dialyzed against Tris-Cl buffer, but was regained after incubating the dialyzed enzyme with dithiothreitol. The regenerated enzyme was inactivated by 5,5-dithiobis(2-nitrobenzoic acid). The resulting inactive thionitrobenzoyl enzyme could be regenerated to the active thiol-enzyme by eithiothreitol or converted to the inactive thiocyanoylated enzyme by KCN. The thiocyanoylated enzyme was insensitive to K⁺ stimulation, which suggested the essentiality of the sulfhydryl groups of theE. coli malic enzyme.     

Regulation of activation of ribulose bisphosphate carboxylase from Pseudomonasoxalaticus

6-phosphogluconate, potentiated activation of ribulose bisphosphate carboxylase from $\text{Pseudomonas}\text{oxalaticus}$ whereas fructose-1,6-bisphosphate inhibited activation and fructose-6-phosphate had no effect. The presence of 1 mM 6-phosphogluconate during activation reduced the K_act for Mg²⁺ from 1.4 mM to approximately 0.2 mM. In the absence of 6-phosphogluconate, the enzyme responded sigmoidally to increasing CO₂ (Hill coefficient, h, of 1.8), with a concentration causing half maximal activation, Act_0.5, of 15 mM NaHCO₃. In the presence of 1 mM 6-phosphogluconate h was reduced to 1.1 and an Act_0.5 value of 5 mM NaHCO₃ was obtained. 6-phosphogluconate appeared to saturate at or below 20 μmM.     

The effect of potassium depletion on the initial kinetics of glycolysis in ascites tumor cells

Ehrlich ascites carcinoma cells depleted of K⁺ and provided with 5.5 mM K⁺ in isosmotic 50 mM tris(hydroxymethyl)methylglycine buffer at pH 7.4 and 38 °C take up K⁺ from the medium at a rate of 6 μmoles/ml intracellular fluid per min. Depleted cells exposed to K⁺ for 2 min prior to glucose addition exhibit a higher initial rate of glycolysis, a lower glycose-6-P accumulation, and a higher fructose-1,6-P₂ accumulation than depleted cells incubated in a K⁺-free medium. Both the K⁺ transport and the effect of K⁺ on glycolysis are blocked by 2 mM oubain.Calculation of thein vitro velocities of glycolytic enzymes from the rates of accumulation of lactate and glycolytic intermediates shows that the presence of K⁺ accelerates the velocities of fructose-6-phosphate kinase and lactate dehydrogenase about 2-fold and the velocity of hexokinase about 1.5-fold during the first 15 s. In either the presence or absence of K⁺, the hexokinase velocity is highest immediately after glucose addition and declines sharply with time; this decline is greater than would be predicted by product inhibition by the accumulated glucose-6-P. The maximal stimulation of fructose-6-phosphate kinase attibutable to the increasing intarcellular K⁺ concentration is only 1.25-fold. These observations indicate that the initial acceleration in glycolysis is not simply mediated through a direct K⁺ activation of fructose-6-phosphate kinase.The calculated theoretical rate of ATP generation by glycolysis shows that glycolysis is an ATP-utilizing system for the first 5–10 s both in the presence and in the absence of K⁺. Hence, the initial stimulation of glycolysis by K⁺ is not a consequence of an increased rate of ATP hydrolysis associated with K⁺ transport, although this mechanism may be responsible for the stimulation of steady-state glycolysis.The initial rate of phosphate ester (hexose and triose phosphates) accumulation corresponds to be rate of ATP generation by the “tail-end” of glycolysis, or twice the rate of lactate accumulation, in either the absence or presence of K⁺, but both the rate and the maximal level of ester accumulated are higher in the presence of K⁺. This implies that the oxidatively generated pool of ATP which is diverted from endogenous reactions to hexokinase and fructose-6-phosphate kinase on the introduction of glucose is larger in the presence of K⁺.Valinomycin (0.27 μM) under certain conditions can produce effects on the glycolysis of non-depleted cells which superficially resemble the effects of K⁺ on depleted cells. However, unlike K⁺, valinomycin stimulates the initial rate of glycolytic ATP generation, and abolishes the initial correspondence between the ATP generation by the “tail-end” of glycolysis and phosphate ester accumulation. These observations are interpreted to mean that valinomycin introduces an ATPase activity effective on glycolytically generated ATP.Comparison of the theoretical ATP generation in the presence and absence of K⁺ indicates that approximately one ATP is hydrolyzed for each K⁺ transported.     

Effects of monovalent cations on phosphate accumulation and storage of the ectomycorrhizal fungus Suillus bovinus

Comparative in vivo ³¹P-NMR studies of the fungus Suillus bovinus (L.: Fr.) O. Kuntze in pure culture have produced interesting new data. To investigate the response of phosphate metabolism to a change in external monovalent cations, samples were exposed to a Hoagland solution containing different monovalent cations Li⁺, Na⁺, K⁺, or Rb⁺ at 10 mM concentration. A method of nutrient cycling during analysis where the cation was changed and the phosphate kept constant allowed us to determine the kinetics of phosphate accumulation, storage and incorporation into polyphosphate following exposure to the range of test cations. Different external monovalent cations had different effects upon changes in the content of both phosphate and polyphosphate. Treatment with Li⁺, Na⁺, or Rb⁺ resulted in a change in phosphate accumulation to 60, 73, and 107% and in content of the intracellular mobile polyphosphate (polyP) to 119, 112, and 94%, respectively, compared with the control taken as 100%. The effect of each cation is related to its position in the periodic table. Reversing this process, i.e., exchanging with K⁺, returned phosphate metabolism to normal. Although, the increase in depolarization of the cell membrane should affect the internal pH, fungal metabolism using energy requiring mechanisms appeared necessary to maintain the intracellular pH. Thus, increasing contents of mobile polyP were the consequence of an increasing energy demand. On the other hand, the increasing depolarization of the cell membrane following the sequence Rb⁺ < K⁺ < Na⁺ < Li⁺ inhibited the net P_i accumulation. Furthermore, it is postulated that the P_i accumulation was also regulated by the intracellular content in polyP.     

pH dependent effects of sodium ions on dextransucrase activity in Streptococcus mutans

Dextransuccrase (E.C 2.4.1.5) is a key enzyme in S. mutans for the metabolism of sucrose which helps in the adherence and accumulation of bacteria on tooth surface leading to the formation of dental caries. Dextransuccrase resembles in its catalytic properties with the brush boarder sucrase and exhibits pH dependent inhibitory and stimulatory effects in response to Na⁺. In this communication we studied the effect of monovalent cations on the activity of dextransuccrase from S. mutans. The percentage inhibition of dextransuccrase was 65% at 0.5 mM NaCl which enhanced to 90% at 20 mM sodium concentration. However there was no effect on dextransucrase activity in presence of other monovalent cations (Rb⁺, Cs⁺, and K⁺) tested. Enzyme activity was enhanced 20–24% in acidic pH but was strongly inhibited (59–89%) around neutral and alkaline pH by 0.5–2.0 mM sodium chloride. Upon dialysis, 86% of enzyme activity was restored to control values. There was no effect of 2 mM NaCl on glucosyltransferase activity of the enzyme. Kinetic studies revealed that enzyme showed biphasic effects in response to Na⁺ ions. At acidic pH the enzyme exhibited mixed type of activation affecting both Vmax and Km, while in alkaline pH, the enzyme showed V- type effect reducing Vmax by 74% without affecting Km. The effects of sodium ions on dextransuccrase activity were specific, thus it can be useful to block its catalytic activity, and reducing the cariogenic potential of S. mutans.     

Comparison of 4-hydroxynzoate decarboxylase and phenol carboxylase activities in cell-free extracts of a defined, 4-hydroxybenzoate and phenol-degrading anaerobic consortium

Summary Two 4-hydroxybenzoate decarboxylase activities and a phenol carboxylase activity were found in cell-free extracts of a defined, 4-hydroxybenzoate- or phenol-grown consortium. Both decarboxylase activities were loosely membrane-associated and required K⁺ but a different pH and ion strength. Loss of activity of both decarboxylases by EDTA could be compensated by Zn²⁺ ions. The K _m values for 4-hydroxybenzoate and K⁺ of the decarboxylase activities with pH optima at 6.4 or 7.8 were 0.02 and 2.5 or 0.004 and 0.5 mm, respectively. 3,4-Dihydroxybenzoate, 3,4,5-tridydroxybenzoate, 3,5-dimethoxy-4-hydroxybenzoate and 3-chloro-4-hydroxybenzoate were also decarboxylated by both enzyme activities. The phenol carboxylase was a soluble enzyme with its pH optimum at 6.5. It required K⁺, Rb⁺ or NH _inf4 ^sup+ as monovalent, Zn²⁺, Mg²⁺, Mn²⁺ or Ni²⁺ as divalent cations and catalysed the carboxylation of phenol if 2,4-,2,3,4- or 2,4,6-hydroxybezoates were absent. The three enzyme activities were not influenced by Avidin and thus were probably not biotin-dependent enzymes. Offprint requests to: J. Winter     

Purification and characterization of an ADP-dependent phosphofructokinase from Thermococcus zilligii

The ADP-dependent phosphofructokinase (PFK) from Thermococcus zilligii has been purified 950 fold; it had a specific activity of 190 U mg⁻¹. The enzyme required Mg²⁺ ions for optimal activity and was specific for ADP. The forward reaction kinetics were hyperbolic for both cosubstrates (pH optimum of 6.4), and the apparent K _m values for ADP and fructose-6-phosphate were 0.6 mM (apparent V _max of 243 U mg⁻¹) and 1.47 mM (apparent V _max of 197 U mg⁻¹), respectively. Significantly, the enzyme is indicated to be nonallosteric but was slightly activated by some monovalent cations including Na⁺ and K⁺. The protein had a subunit size of 42.2 kDa and an estimated native molecular weight of 66 kDa (gel filtration). Maximal reaction rates for the reverse reaction were attained at pH 7.5–8.0, and the apparent K _m values for fructose-1,6-bisphosphate and AMP were 0.56 mM (apparent V _max of 2.9 U mg⁻¹) and 12.5 mM, respectively. The biochemical characteristics of this unique ADP-dependent enzymatic activity are compared to ATP and pyrophosphate-dependent phosphofructokinases. Received: August 14, 1998 / Accepted: December 2, 1998     

The role of monovalent cations for photosynthesis of isolated intact chloroplasts

The role of monovalent cations in the photosynthesis of isolated intact spinach chloroplasts was investigated. When intact chloroplasts were assayed in a medium containing only low concentrations of mono- and divalent cations (about 3 mval l^-1), CO₂-fixation was strongly inhibited although the intactness of chloroplasts remained unchanged. Addition of K⁺, Rb⁺, or Na⁺ (50–100 mM) fully restored photosynthesis. Both the degree of inhibition and restoration varied with the plant material and the storage time of the chloroplasts in low-salt medium. In most experiments the various monovalent cations showed a different effectiveness in restoring photosynthesis of low-salt chloroplasts (K⁺>Rb⁺>Na⁺). Of the divalent cations tested, Mg²⁺ also restored photosynthesis, but to a lesser extent than the monovalent cations.In contrast to CO₂-fixation, reduction of 3-phosphoglycerate was not ihibited under low-salt conditions. In the dark, CO₂-fixation of lysed chloroplasts supplied with ATP, NADPH, and 3-phosphoglycerate strictly required the presence of Mg²⁺ but was independent of monovalent cations. This finding excludes a direct inactivation of Calvin cycle enzymes as a possible basis for the inhibition of photosynthesis under low-salt conditions.Light-induced alkalization of the stroma and an increase in the concentration of freely exchangeable Mg²⁺ in the stroma, which can be observed in normal chloroplasts, did not occur under low-salt conditions but were strongly enhanced after addition of monovalent cations (50–100 mM) or Mg²⁺ (20–50 mM).The relevance of a light-triggered K⁺/H⁺ exchange at the chloroplast envelope is discussed with regard to the light-induced increase in the pH and the Mg²⁺ concentration in the stroma, which are thought to be obligatory for light activation of Calvincycle enzymes.     

Activation of diol dehydrase by formamidinium or guanidinium ion, polyatomic monovalent cations having sp2 nitrogen

Coenzyme B₁₂-dependent diol dehydrase was activated by formamidinium or guanidinium ion. These polyatomic monovalent cations having sp² hybrid atomic orbitals and trigonal orientation were much more effective in activating the enzyme than methylammonium ion, but less active than NH₄+ or K⁺. Formamidinium and guanidinium ions were also effective both in forming and maintaining the binding of coenzyme B₁₂ to the apoenzyme. There is a close relationship between the effectiveness in activating the enzyme and those in forming and maintaining the holoenzyme, suggesting that these polyatomic monovalent cations play the same role in the diol dehydrase system as alkali metal monovalent cation such as K⁺.     

Ion complexation in nonactin,monactin, and dinactin: A Raman spectroscopic study

The ability of the macrotetrolide nactins to complex selectivity with a wide variety of cations makes these ionophorous antibiotics important model systems for the study of biologic ionic transport. We report a Raman spectroscopic investigation of the Na⁺, K⁺, Rb⁺, Cs⁺, Tl⁺, NH₄⁺, NH₃OH⁺, C(NH₂)₃⁺, and Ba⁺⁺ complexes of nonactin, monactin, and dinactin in 4:1 (v/v) CH₃OH/CHCl₃ and in the solid state. The nactins display characteristic spectral changes upon complexation, some of which are specific for a given cation. In the K⁺, Rb⁺, Cs⁺, NH₃OH⁺, and C(NH₂)₃⁺ complexes, which are apparently isosteric, the ester carbonyl stretch frequency is found to be linearly proportional to the cation–carbonyl electrostatic interaction energy, as calculated from a simplified model. Deviations for the Na⁺, NH₄⁺, Tl⁺, and Ba⁺⁺ complexes are interpreted as arising from additional nonelectrostatic interactions. Additional information is obtained from other spectral regions and from measurements of depolarization ratios. Spectra of the nactin complexes differ from each other more in the solid state than in solution, reflecting the effects of crystalline contact forces.     

Ion conduction and selectivity in acid-sensing ion channel 1

The ability of acid-sensing ion channels (ASICs) to discriminate among cations was assessed based on changes in conductance and reversal potential with ion substitution. Human ASIC1a was expressed in Xenopus laevis oocytes, and acid-induced currents were measured using two-electrode voltage clamp. Replacement of extracellular Na⁺ with Li⁺, K⁺, Rb⁺, or Cs⁺ altered inward conductance and shifted the reversal potentials consistent with a selectivity sequence of Li ∼ Na > K > Rb > Cs. Permeability decreased more rapidly than conductance as a function of atomic size, with P_K/P_Na = 0.1 and G_K/G_Na = 0.7 and P_Rb/P_Na = 0.03 and G_Rb/G_Na = 0.3. Stimulation of Cl⁻ currents when Na⁺ was replaced with Ca²⁺, Sr²⁺, or Ba²⁺ indicated a finite permeability to divalent cations. Inward conductance increased with extracellular Na⁺ in a hyperbolic manner, consistent with an apparent affinity (K_m) for Na⁺ conduction of 25 mM. Nitrogen-containing cations, including NH₄⁺, NH₃OH⁺, and guanidinium, were also permeant. In addition to passing through the channels, guanidinium blocked Na⁺ currents, implying competition for a site within the pore. The role of negative charges in an external vestibule of the pore was evaluated using the point mutation D434N. The mutant channel had a decreased single-channel conductance, measured in excised outside-out patches, and a macroscopic slope conductance that increased with hyperpolarization. It had a weakened interaction with Na⁺ (K_m = 72 mM) and a selectivity that was shifted toward larger atomic sizes. We conclude that the selectivity of ASIC1 is based at least in part on interactions with binding sites both within and internal to the outer vestibule.     

Ability of monovalent cations to replace potassium during stimulation of lymphoid cells

Stimulation of hamster thymocytes, splenocytes, or lymph node cells occurred to a minimal extent in the absence of K⁺. This observation was found for stimulation by T-cell mitogens (phytohemagglutinin and concanavalin A), A B-cell mitogen (lipopolysaccharide), or antigen (KLH). Marginal restoration of the responses to these stimulants occurred in the presence of 0.1 mM K⁺ and responsiveness returned to near maximal levels on addition of 1 mM K⁺ to the cultures. Attempts to restore the responsiveness with other monovalent cations revealed an order of effectiveness of K⁺ ≥ Rb⁺ ? NH₄⁺ ≥ Li⁺. At the 1 mM level K⁺ and Rb⁺ were equally effective in supporting stimulation by phytohemagglutinin while all concentrations of Li⁺ tested (0.1–10 mM) would not support stimulation. However, addition of Li⁺ to cultures reconstituted with 1 mM K⁺ or Rb⁺ revealed that this ion could enhance the phytohemagglutinin response by approximately 100% in the presence of K⁺ and only 30% in the presence of Rb⁺. These data support the hypotheses that the Na,K ATPase must be active for lymphocyte stimulation to occur and that some of the biological effects of Li⁺ on lymphocyte stimulation are mediated at the level of the Na,K ATPase.     

Activation of wheat germ acetyl CoA carboxylase by potassium and rubidium.

Wheat germ acetyl CoA carboxylase requires certain alkali cations to exhibit maximal activity. Maximal activation results when 60 mM K⁺ or Rb⁺ are included in the assay mixture, whereas only marginal activation occurs in the presence of similar concentrations of Li⁺⁺ and Na⁺⁺. Cs⁺⁺ activates, but less effectively than K⁺ or Rb⁺. Since it is also possible to activate the enzyme maximally using 20 mM potassium isocitrate, but not 20 mM sodium isocitrate, activation of the wheat germ enzyme is due to a cation effect and not to citrate anion.