Effect of lithium salts on lactate dehydrogenase,adenylate kinase,and 1-phosphofructokinase activities

Inhibitions of 30?nM rabbit muscle 1-phosphofructokinase (PFK-1) by lithium, potassium, and sodium salts showed inhibition or not depending upon the anion present. Generally, potassium salts were more potent inhibitors than sodium salts; the extent of inhibition by lithium salts also varied with the anion. Li₂CO₃ was a relatively potent inhibitor of PFK-1 but LiCl and lithium acetate were not. Our results suggest that extents of inhibition by monovalent salts were due to both cations and anions, and the latter needs to be considered before inhibition can be credited to the cation. An explanation for monovalent salt inhibitions is proffered involving interactions of both cations and anions at negative and positive sites of PFK-1 that affect enzyme activity. Our studies suggest that lithium cations per se are not inhibitors: the inhibitors are the lithium salts, and we suggest that in vitro studies involving the effects of monovalent salts on enzymes should involve more than one anion.     

Properties of pyruvate kinase from soybean nodule cytosol

The properties of pyruvate kinase from soybean (Glycine max L.) nodule cytosol were examined to determine what influence the N₂ fixation process might have on this supposed key control enzyme. A crude enzyme preparation was prepared by chromatography of cytosol extract on a diethylaminoethyl-cellulose column. ATP and citrate at 5 mm concentrations inhibited pyruvate kinase 27 and 34%, respectively. Enzyme activation was hyperbolic with respect to both K⁺ and NH₄⁺ concentrations. In the presence of physiological concentrations of K⁺ and high phosphoenolpyruvate (PEP) concentrations, NH₄⁺ inhibited enzyme activity. Comparisons of kinetic parameters (V_max and apparent K_a) for NH₄⁺ and K⁺ with inhibition curves indicated that inhibition was very likely a result of competition of the ions for activation site(s) on the pyruvate kinase. In addition, apparent K_a (monovalent cation) and K_m (PEP) were influenced by PEP and monovalent cation concentrations, respectively. This effect may reflect a fundamental difference between plant and animal pyruvate kinases. It is concluded that control of cytosol pyruvate kinase may be closely related to reactions involved in the assimilation of NH₄⁺.     

Some properties of pyruvate kinase extracted from Lycopersicon esculentum

Pyruvate kinase was extracted from Me₂CO-dried tissue of various parts of tomato plants. Recovery of the enzyme was improved by the inclusion of thiols in the extraction medium, and its stability was increased considerably in the presence of glycerol and to a lesser extent tetramethylammonium chloride. A phosphatase was present in the tissue extracts which hydrolyses phosphoenolpyruvate in the absence of added ADP. ATP inhibited pyruvate kinase but stimulated the phosphatase, while Mg²⁺ stimulated both enzymes. Data obtained suggest that tomato leaf pyruvate kinase has an absolute dependence on monovalent cations for activity, K⁺ being the principal activator. The phosphatase was inhibited non-selectively by monovalent cations. The total activity of pyruvate kinase and its concentration on a tissue fresh weight basis was greatest in the leaves, activity increasing with the maturity of the tissue. Less enzyme was present in roots, and least in the fruit.     

Some characteristics of Rabbit muscle phosphofructokinase-1 inhibition by ascorbate

These studies relate to a working hypothesis that glycogen storage is facilitated in resting muscle by inhibiting glycolysis via inhibition of LDH, AK, and PFK-1 by ascorbate; when muscle is active, these isozymes combine with muscle proteins and are released and protected from inhibition by ascorbate and glycolysis proceeds. Focus in these studies is on the ability of G-actin and aldolase to prevent PFK-1 inhibition by ascorbate. We found that inhibition by ascorbate was PFK-1 concentration dependent; ascorbate does not inhibit above 200 nM PFK-1. We conclude that ascorbate inhibits PFK-1 dimers (and perhaps monomers) but not PFK-1 tetramers. Separation of PFK-1 dimers from tetramers was achieved with centrifugal filter devices and differences in their sensitivity to ascorbate inhibition were demonstrated. Some comparisons are made with attributes of AK inhibitions by ascorbate that, like PFK-1, are also enzyme concentration dependent. Discussions relate findings to cellular infrastructure and the role of ascorbate in glycogen synthesis.     

Activation of an ATP-dependent K+ conductance in Xenopus oocytes by expression of adenylate kinase cloned from renal proximal tubules

In rabbit proximal convoluted tubules, an ATP-sensitive K⁺ (K_ATP) channel has been shown to be involved in membrane cross-talk, i.e. the coupling (most likely mediated through intracellular ATP) between transepithelial Na⁺ transport and basolateral K⁺ conductance. This K⁺ conductance is inhibited by taurine. We sought to isolate this K⁺ channel by expression cloning in Xenopus oocytes. Injection of renal cortex mRNA into oocytes induced a K⁺ conductance, largely inhibited by extracellular Ba²⁺ and intracellular taurine. Using this functional test, we isolated from our proximal tubule cDNA library a unique clone, which induced a large K⁺ current which was Ba²⁺-, taurine- and glibenclamide-sensitive. Surprisingly, this clone is not a K⁺ channel but an adenylate kinase protein (AK3), known to convert NTP+AMP into NDP+ADP (N could be G, I or A). AK3 expression resulted in a large ATP decrease and activation of the whole-cell currents including a previously unknown, endogenous K⁺ current. To verify whether ATP decrease was responsible for the current activation, we demonstrated that inhibition of glycolysis greatly reduces oocyte ATP levels and increases an inwardly rectifying K⁺ current. The possible involvement of AK in the K_ATP channel’s regulation provides a means of explaining their observed activity in cytosolic environments characterized by high ATP concentrations.     

Purification and Characterization of a Cation-stimulated Adenosine Triphosphatase from Corn Roots

A membrane-bound, monovalent cation-stimulated ATPase from Zea mays roots has been purified to a single band on sodium dodecyl sulfate gel electrophoresis. Microsomal preparations with K⁺ -stimulated ATPase activity were extracted with 1 m NaClO₄, and the solubilized enzyme was purified by chromatography on columns of n-hexyl-Sepharose, DEAE-cellulose, and Sephadex G-100 Superfine. A 500-fold purification over the activity present in the microsomes was obtained. The K⁺ -stimulated activity shows positive cooperativity with increasing KCl concentrations. The purified enzyme shows K⁺ -stimulated activity with ATP, GTP, UTP, CTP, ADP, α + β-glycerophosphate, p-nitrophenyl phosphate, and pyrophosphate as substrates. Under most conditions ATP is the best substrate. Although dicyclohexyl carbodiimide and Ca²⁺ inhibit and alkylguanidines stimulate the K⁺ -ATPase while bound to microsomes, they have no effect on the purified enzyme.     

Correlation between monovalent cation-induced decreases in chlorophyll a fluorescence and chloroplast structural changes

Low concentrations (~ 3 mm) of salts of monovalent cations such as Na⁺, K⁺, and tetraethylammonium were found to decrease the turbidity of chloroplast suspensions. The turbidity changes (Δ₅₄₀) had the same kinetics, salt concentration dependence, and pH dependence as the monovalent cation-induced decreases in chlorophyll a fluorescence (9), suggesting that structural changes are the cause of the associated increases in spillover. Electron microscopy revealed that the grana are stacked when spillover is inhibited (in the absence of salts or the presence of divalent cations) and that monovalent cations cause the grana to unstack, thereby promoting spillover.     

Changes in growth and water-soluble solute concentrations in Sorghum bicolor stressed with sodium and potassium salts

Sorghum bicolor L. Moench, RS 610, was grown in liquid media salinized with NaCl, KCl, Na₂SO₄, K₂SO₄ or with variable mixtures of either NaCl/KCl or Na₂SO₄/K₂SO₄ at osmotic potentials ranging from 0 to -0.8 MPa. The purpose was to study the effects of different types and degrees of salinity in growth media on growth and solute accumulation. In 14-day-old plants the severity of leaf growth inhibition at any one level of osmotic potential in the medium increased according to the following order: NaCl < Na₂SO₄ < KCl = K₂SO₄. Inhibition of growth by mixtures of Na⁺ and K⁺ salts was the same as by K⁺ salts alone. Roots responded differently. Root growth was not affected by Na⁺ salts in the range of 0 to -0.2 MPa while it was stimulated by K⁺ salts. The major cation of leaves was K⁺ because S. bicolor is a Na⁺-excluder, while Na⁺ was the major cation in roots except at low Na⁺/K⁺ ratios in media. Anions increased in tissues linearly in relation to total monovalent cation, but not with a constant anion/cation ratio. This ratio increased as the cation concentrations in tissues increased. Sucrose in leaf tissue increased 75 fold in Chloride-plants (plants growing in media in which the only anion of the salinizing salts was Cl^?) and 50 fold in Sulphate-plants (the only anion of the salinizing salts was SO₄^2-). Proline increased 60 and 18 fold in Chloride- and Sulphate-plants, respectively, as growth media potentials decreased from 0 to -0.8 MPa. The concentrations of both sucrose and proline were directly proportional to the amount of total monovalent cation in the tissue. Sucrose concentrations began increasing when total monovalent cations exceeded 100 μmol (g fresh weight)^?1 (the monovalent cation level in non-stressed plants), but proline did not start accumulating until monovalent cation concentrations exceeded 200 μmol (g fresh weight)^?1. Therefore, sucrose seemed to be the solute used for osmotic adjustment under mild conditions of saline stress while proline was involved in osmotic adjustment under more severe conditions of stress. Concentrations of inorganic phosphate, glucose, fructose, total amino acids and malic acid fluctuated in both roots and leaves in patterns that could be somewhat correlated with saline stress and, sometimes, with particular salts in growth media. However, the changes measured were too small (at most a 2–3 fold increase) to be of importance in osmotic adjustment.     

Monovalent cation requirement for ADP inhibition of pyruvate dehydrogenase kinase

ADP is a competitive inhibitor with respect to ATP for pyruvate dehydrogenase kinase. Evidence is presented that K⁺ or NH₄⁺ ions are required for inhibition of the kinase by ADP. K⁺ at 30–90 mM and NH₄⁺ at 1–5 mM decrease markedly the apparent K_i of bovine kidney pyruvate dehydrogenase kinase for ADP and also decrease, to a lesser extent, the apparent K_m for ATP. Na⁺ is less effective and, in addition, inhibits kinase activity. Since K⁺ and NH₄⁺ are not required for kinase activity, their effect appears to be primarily of regulatory significance. K⁺ and NH₄⁺ have little effect, if any, on pyruvate dehydrogenase phosphatase activity. When both the kinase and the phosphatase are present and functional, the near steady state activity of the pyruvate dehydrogenase complex is affected significantly by varying the concentration of K⁺ or NH₄⁺ at a fixed ADP/ATP concentration ratio and by varying the $\text{ADP}\text{ATP}$ ratio at a fixed concentration of monovalent cation.     

Modification of foliar solute concentrations by calcium in two species of wheat stressed with sodium chloride and/or potassium chloride

The effects of saline-stresses due to different salts on growth and on foliar solute concentrations in seedlings of two species of wheat that differed in salt tolerance. Triticum aestivum L. cv. Probred and Triticum turgidum L. (Durum group) cv. Aldura, were studied. Triticum aestivum is the more salt tolerant species. The salts used were NaCl, KCI, a 1:1 mixture of NaCI and KCI, and these same monovalent cation salts but mixed with CaCI₂ at a ratio of 2:1 on a molar basis of monovalent to divalent cation salts. Growth inhibition of both species was a function of media osmotic potentials. There was a small additional inhibition of growth if KCI replaced NaCI as the salinizing salt. CaCI₂ had little or no effect on growth inhibition beyond an osmotic effect except at the most severe stress level, i.e. when Ca²⁺ concentrations may be excessive. The amounts of water-soluble Ca²⁺ were about 10 times higher in leaves of plants grown in the presence of CaCI₂ than in its absence, but its concentrations even then were approximately 10% or less of those of the monovalent cations. Including CaCI₂ in growth media resulted in a reduction in the amount of Na⁺ in leaves compared to the amounts in plants grown at the same osmotic potential but in the absence of CaCI₂. Triticum aestivum was a better Na⁺-excluder than T. turgidum. With CaCI₂ in media, (Na⁺+ K⁺) remained relatively constant or increased by small amounts as media osmotic potentials décreased. In the absence of CaCI₂₊ (Na⁺+ K⁺) increased by large amounts when media osmotic potentials were at ?0.6 and ?0.8 MPa. It is concluded that the accumulation system in leaves for monovalent cations was under feed-back control, and that this control mechanism was inhibited by high media concentrations of Na⁺ and/or K⁺. Sucrose was present at a constant amount under all growth conditions. Proline started accumulating when (Na⁺+ K⁺) exceeded a threshold value of 200 μmol (g fresh weight)^?1. Its concentration was 5 to 13% of that portion of (Na⁺+ K⁺) that exceeded the threshold value.     

Impact of inorganic salts on degradation of bisphenol A and diclofenac by crude extracellular enzyme from Pleurotus ostreatus

This study investigated the influence of inorganic salts on enzymatic activity and the removal of trace organic contaminants (TrOCs) by crude laccase from the white-rot fungus Pleurotus ostreatus. A systematic analysis of 15 cations and anions from common inorganic salts was presented. Laccase activity was not inhibited by monovalent cations (i.e. Na⁺, NH₄⁺, K⁺), while the presence of divalent and trivalent cations showed variable impact – from negligible to complete inhibition – of both laccase activity and its TrOC removal performance. Of interest was the observation of discrepancy between residual laccase activity and TrOC removal in the presence of some ions. Mg²⁺ had negligible impact on residual laccase activity but significant impact on TrOC removal. Conversely, F^? showed greater impact on residual laccase activity than on TrOC removal. This observation indicated different impacts of the interfering ions on the interaction between laccase and TrOCs as compared to that between laccase and the reagent used to measure its activity, implicating that residual laccase activity may not always be an accurate indicator of TrOC removal. The degree of impact of halides was in the order of F^??>?I^? >?Br^??>?Cl^?. Particularly, the tolerance of the tested laccase to Cl^? has important implications for a range of industrial applications.     

Characterization of a partially purified adenosine triphosphatase from a corn root plasma membrane fraction

The (K⁺,Mg²⁺)-ATPase was partially purified from a plasma membrane fraction from corn roots (WF9 × Mol7) and stored in liquid N₂ without loss of activity. Specific activity was increased 4-fold over that of the plasma membrane fraction. ATPase activity resembled that of the plasma membrane fraction with certain alterations in cation sensitivity. The enzyme required a divalent cation for activity (Co²⁺ > Mg²⁺ > Mn²⁺ > Zn²⁺ > Ca²⁺) when assayed at 3 millimolar ATP and 3 millimolar divalent cation at pH 6.3. When assayed in the presence of 3 millimolar Mg²⁺, the enzyme was further activated by monovalent cations (K⁺, NH₄⁺, Rb⁺ Na⁺, Cs⁺, Li⁺). The pH optima were 6.5 and 6.3 in the absence and presence of 50 millimolar KCl, respectively. The enzyme showed simple Michaelis-Menten kinetics for the substrate ATP-Mg, with a K_m of 1.3 millimolar in the absence and 0.7 millimolar in the presence of 50 millimolar KCl. Stimulation by K⁺ approached simple Michaelis-Menten kinetics, with a K_m of approximately 4 millimolar KCl. ATPase activity was inhibited by sodium orthovanadate. Half-maximal inhibition was at 150 and 35 micromolar in the absence and presence of 50 millimolar KCl. The enzyme required the substrate ATP. The rate of hydrolysis of other substrates, except UDP, IDP, and GDP, was less than 20% of ATP hydrolysis. Nucleoside diphosphatase activity was less than 30% of ATPase activity, was not inhibited by vanadate, was not stimulated by K⁺, and preferred Mn²⁺ to Mg²⁺. The results demonstrate that the (K⁺,Mg²⁺)-ATPase can be clearly distinguished from nonspecific phosphohydrolase and nucleoside diphosphatase activities of plasma membrane fractions prepared from corn roots.     

Effects of monovalent cations on the catalytic activity of pig liver phosphofructokinase.

The monovalent cations NH₄⁺, K⁺, and Rb⁺ activate pig liver phosphofructokinase by increasing the maximal velocity. In the presence of these cations the enzyme retains sigmoid kinetics with respect to fructose-6-phosphate. However, these cations bring about a decrease in the [S]_0.5 for fructose-6-phosphate to an extent directly proportional to their ionic volumes. The apparent dissociation constants of NH₄⁺, K⁺, and Rb⁺ for the enzyme at 0.5 mM ATP and 4 mM Fru6P are 0.2 mM, 8 mM, and 15 mM, respectively. The maximal velocity of the enzyme in the presence of saturating concentrations of Rb⁺ is about 70% of that seen with NH₄⁺ or K⁺. The monovalent cations Li⁺, Na⁺, and Cs⁺ inhibit the enzyme at high concentrations (> 50 mM) by decreasing the maximal velocity. Although the efficiency of inhibition by these cations qualitatively increases with decreasing size, there is no obvious quantitative relationship between efficiency of inhibition and any parameter of ionic size.     

Effects of Cations on (Ca2++ Mg2+)-Activated ATPase from Rat Brain

Abstract: With a partially purified, membrane-bound (Ca + Mg)-activated ATPase preparation from rat brain, the K_0.5 for activation by Ca²⁺ was 0.8 p μm in the presence of 3 mm -ATP, 6 mm -MgCl₂, 100 m_M-KCI, and a calcium EGTA buffer system. Optimal ATPase activity under these circumstances was with 6-100 μm -Ca²⁺, but marked inhibition occurred at higher concentrations. Free Mg²⁺ increased ATPase activity, with an estimated K_0.5, in the presence of 100 μm -CaCl₂, of 2.5 mm ; raising the MgCl₂ concentration diminished the inhibition due to millimolar concentrations of CaCl₂, but antagonized activation by submicromolar concentrations of Ca²⁺. Dimethylsulfoxide (10%, v/v) had no effect on the K_0.5 for activation by Ca²⁺, but decreased activation by free Mg²⁺ and increased the inhibition by millimolar CaCl₂. The monovalent cations K⁺, Na⁺, and TI⁺ stimulated ATPase activity; for K⁺ the K_0.5 was 8 mm , which was increased to 15 mm in the presence of dimethylsulfoxide. KCI did not affect the apparent affinity for Ca²⁺ as either activator or inhibitor. The preparation can be phosphorylated at 0°C by [γ-³²P]-ATP; on subsequent addition of a large excess of unlabeled ATP the calcium dependent level of phosphorylation declined, with a first-order rate constant of 0.12 s^?1. Adding 10 mm -KCI with the unlabeled ATP increased the rate constant to 0.20 s^?1, whereas adding 10 mm -NaCl did not affect it measurably. On the other hand, adding dimethyl-sulfoxide slowed the rate of loss, the constant decreasing to 0.06 s^?1. Orthovanadate was a potent inhibitor of this enzyme, and inhibition with 1 μm -vanadate was increased by both KCI and dimethylsulfoxide. Properties of the enzyme are thus reminiscent of the plasma membrane (Na + K)-ATPase and the sarcoplasmic reticulum (Ca + Mg)-ATPase, most notably in the K⁺ stimulation of both dephosphorylation and inhibition by vanadate.     

5-Oxoprolinase (l-Pyroglutamate Hydrolase) in Higher Plants: Partial Purification and Characterization of the Wheat Germ Enzyme

5-Oxoprolinase has been found to be widely distributed in higher plants. This enzyme catalyzes the ATP-dependent hydrolysis of 5-oxo-l-proline (l-pyrollidone carboxylate, l-pyroglutamate) to glutamate. The enzyme has been purified almost 60 fold from wheat germ (Triticum aestivum L). This enzyme requires a divalent cation, either Mn²⁺ or Mg²⁺, and a combination of both appears to be the most effective. There is also an absolute requirement for a monovalent cation best fulfilled by either NH₄⁺ or K⁺. The K_m for ATP is 0.4 mm and for 5-oxo-l-proline is 14 μm. A small amount of activity is observed when other purine nucleotides such as ITP and GTP replace ATP. The substitution of the pyrimidine nucleotides CTP and UTP for ATP yield almost completely inactive preparations. The enzyme appears to have an active sulfhydryl group since there is an increase in activity in the presence of dithioerythritol. Preincubation with reagents such as N-ethylmaleimide or iodoacetamide lead to complete inactivation. The presence of this enzyme leads to the speculation of the possible presence of a γ-glutamyl cycle in higher plants.     

Some characteristics of rabbit muscle phosphofructokinase-1 inhibition by ascorbate

These studies relate to a working hypothesis that glycogen storage is facilitated in resting muscle by inhibiting glycolysis via inhibition of LDH, AK, and PFK-1 by ascorbate; when muscle is active, these isozymes combine with muscle proteins and are released and protected from inhibition by ascorbate and glycolysis proceeds. Focus in these studies is on the ability of G-actin and aldolase to prevent PFK-1 inhibition by ascorbate. We found that inhibition by ascorbate was PFK-1 concentration dependent; ascorbate does not inhibit above 200 nM PFK-1. We conclude that ascorbate inhibits PFK-1 dimers (and perhaps monomers) but not PFK-1 tetramers. Separation of PFK-1 dimers from tetramers was achieved with centrifugal filter devices and differences in their sensitivity to ascorbate inhibition were demonstrated. Some comparisons are made with attributes of AK inhibitions by ascorbate that, like PFK-1, are also enzyme concentration dependent. Discussions relate findings to cellular infrastructure and the role of ascorbate in glycogen synthesis.     

Schistosoma mansoni: effect of some cations on the proteolytic enzymes of cercariae

The effects of various salts on the proteolytic activity of extracts from Schistosoma mansoni cercariae were tested. Using an Azocoll substrate, stimulation (2 to 2.5-fold) of activity by the monovalent cations Na⁺ and K⁺ was demonstrated, with maximum stimulation at 20–40 mM concentrations. The divalent cations Mg²⁺ and Ca²⁺ stimulated proteolytic activity at low concentrations (between 0 and 10 mM) but inhibited activity at higher concentrations. The divalent cations Zn²⁺, Cu²⁺, Fe²⁺, and Co²⁺ were inhibitory even at very low concentrations. The results presented here are discussed in relation to previously described ion effects on cercarial infectivity.     

AMPK modulates glucose-sensing in insulin-secreting cells by altered phosphotransfer to KATP channels

Glucose-sensing (GS) behaviour in pancreatic β-cells is dependent on ATP-sensitive K⁺ channel (K_ATP) activity, which is controlled by the relative levels of the K_ATP ligands ATP and ADP, responsible for closing and opening K_ATP, respectively. However, the mechanism by which β-cells transfer energy status from mitochondria to K_ATP, and hence to altered electrical excitability and insulin secretion, is presently unclear. Recent work has demonstrated a critical role for AMP-activated protein kinase (AMPK) in GS behaviour of cells. Electrophysiological recordings, coupled with measurements of gene and protein expression were made from rat insulinoma cells to investigate whether AMPK activity regulates this energy transfer process. Using the whole-cell recording configuration with sufficient intracellular ATP to keep K_ATP closed, raised AMPK activity induced GS electrical behaviour. This effect was prevented by the AMPK inhibitor, compound C and required a phosphotransfer process. Indeed, high levels of intracellular phosphocreatine or the presence of the adenylate kinase (AK) inhibitor AP₅A blocked this action of AMPK. Using conditions that maximised AMPK-induced K_ATP opening, there was a significant increase in AK1, AK2 and UCP2 mRNA expression. Thus we propose that K_ATP opening in response to lowered glucose concentration requires AMPK activity, perhaps in concert with increased AK and UCP2 to enable mitochondrial-derived ADP signals to be transferred to plasma membrane K_ATP by phosphotransfer cascades.     

Adenosine Triphosphatases Bound to Plasmalemma, Mitochondria, and Microsomes or Present in the Cytoplasmic Phase from Potato Tubers

Between pH 4–10, basal ATPase activity, measured in the absence of mineral ions, was 10 to 100 times higher in the final cytoplasmic supernatant from potato tuber homogenates than in the membraneous fractions (purified plasmalemma, purified mitochondria and microsomes). The soluble ATPase was slightly inhibited, whereas the membrane-bound ATPases were all stimulated by Mg²⁺ ions. A further stimulation by Na⁺ or K⁺ ions was only observed in purified plasmalemma or mitochondria, at alkaline pH (7.5–9.5). At a fixed (Na⁺+ K⁺) concentrations (80 mM), this last stimulation was much greater in purified mitochondria (350%) than in plasmalemma (33%); it also increased with (Na⁺+ K⁺) concentrations up to 200 mM in mitochondria whereas, in plasmalemma, it was roughly constant for monovalent ion concentrations between 20 and 200 mM. General properties of the plasma membrane-bound ATPase have been determined, i.e. substrate specificity, activity variations with quantity of substrate, temperature, pH, etc. Divalent cations stimulated strongly the ATPase in the following order: Mn²⁺ > Mg²⁺ > Ca²⁺. The maximum ATP hydrolysis velocity for that part of ATPase activity which is strictly dependent on Mg²⁺ ions was 3.85 μmol × mg^?1 protein × h^?1. This plasma membrane ATPase was not sensitive to ouabaïn or to oligomycin.