scholarly journals Role of the subunits of the energy-transducing adenosine triphosphatase from Micrococcus lysodeikticus membranes studied by proteolytic digestion and immunological approaches

1980 ◽  
Vol 186 (3) ◽  
pp. 713-723 ◽  
Author(s):  
F Mollinedo ◽  
V Larraga ◽  
F J Coll ◽  
E Muñoz
Keyword(s):  
Atpase Activity ◽  
Adenosine Triphosphatase ◽  
Specific Activity ◽  
Relative Proportion ◽  
Inhibitory Effect ◽  
Beta Subunit ◽  
Gamma Subunit ◽  
Micrococcus Lysodeikticus ◽  
Alpha Beta

An energy-transducing adenosine triphosphatase (ATPase, EC 3.6.1.3) that contains an extra polypeptide (delta) as well as three intrinsic subunits (alpha, beta, gamma) was purified from Micrococcus lysodeikticus membranes. The apparent subunit stoichiometry of this soluble ATPase complex is alpha 3 beta 3 gamma delta. The functional role of the subunits was studied by correlating subunit sensitivity to trypsin and effect of antibodies raised against holo-ATPase and its alpha, beta and gamma subunits with changes in ATPase activity and ATPase rebinding to membranes. A form of the ATPase with the subunit proportions 1.67(alpha):3.00(beta:0.17(gamma) was isolated after trypsin treatment of purified ATPase. This form has more than twice the specific activity of native enzyme. Other forms with less relative proportion of alpha subunits and absence of gamma subunit are not active. Of the antisera to subunits, only anti-(beta-subunit) serum shows a slight inhibitory effect on ATPase activity, but its combination with either anti-(alpha-subunit) or anti-(gamma-subunit) serum increases the effect. The results suggest that beta subunit is required for full ATPase activity, although a minor proportion of alpha and perhaps gamma subunit(s) is also required, probably to impart an active conformation to the protein. The additional polypeptide not hitherto described in Micrococcus lysodeikticus ATPase had a molecular weight of 20 000 and was found to be involved in ATPase binding to membranes. This 20 000-dalton component can be equated with the delta subunit of other energy-transducing ATPases and its association with the (alpha, beta, gamma) M. lysodeikticus ATPase complex appears to be dependent on bivalent cations. The present results do not preclude the possibility that the gamma subunit also plays a role in ATPase binding, in which, however, the major subunits do not seem to play a role.

Mercury weakens membrane anchoring of Na-K-ATPase

1992 ◽  
Vol 262 (5) ◽  
pp. F837-F842 ◽  
Author(s):  
E. Imesch ◽  
M. Moosmayer ◽  
B. M. Anner
Keyword(s):  
Atpase Activity ◽  
Model Compound ◽  
Beta Subunit ◽  
Alpha Subunit ◽  
Structural Effects ◽  
Subunit Interaction ◽  
Alpha Beta ◽  
Membrane Anchoring ◽  
Atpase Inhibition

The presence of circulating inhibitors able to decrease the renal Na-K-adenosinetriphosphatase (ATPase) activity (natriuretic hormones) was postulated some 30 years ago. In the present work, the natriuretic inhibitor HgCl2 was selected as a model compound for the structural characterization of a possible natriuretic pathway for Na-K-ATPase modification. The structural effects of Na-K-ATPase inhibition by HgCl2 were assessed by trypsinolysis of the blocked enzyme in comparison with untreated preparations. The results show that inactivation of Na-K-ATPase by HgCl2 leads to the release of the alpha-subunit from the membrane preferentially in the E2 conformation but also in the E1 conformation. Apparently, HgCl2 weakens the membrane anchoring of the alpha-subunit, presumably by loosening the alpha-beta-subunit interaction. By this mechanism, the sensitivity of the Na-K-ATPase to extracellular drugs, hormones, and antibodies, as well as to intracellular proteases and other regulatory factors, could be altered.

scholarly journals The effect of the γ-subunit of the cyclic GMP phosphodiesterase of bovine and frog (Rana catesbiana) retinal rod outer segments on the kinetic parameters of the enzyme

1990 ◽  
Vol 265 (3) ◽  
pp. 655-658 ◽  
Author(s):  
M M Whalen ◽  
M W Bitensky ◽  
D J Takemoto
Keyword(s):  
Catalytic Activity ◽  
Cyclic Gmp ◽  
Active Sites ◽  
Outer Segment ◽  
Maximal Activity ◽  
Inhibitory Effect ◽  
Gamma Subunit ◽  
Rod Outer Segment ◽  
Gamma Subunits ◽  
Alpha Beta

Rod-outer-segment cyclic GMP phosphodiesterase (PDE) (subunit composition alpha beta gamma 2) contains catalytic activity in alpha beta. The gamma-subunits are inhibitors. Removal of the gamma-subunits increases Vmax. without affecting the Km. The inhibitory effect of a single gamma-subunit (alpha beta gamma) on the Vmax. of alpha beta is much greater in bovine than in frog (Rana catesbiana) PDE. Bovine PDE in the alpha beta gamma 2 state has a Vmax. that is 2.6 +/- 0.4% of the Vmax. of alpha beta. The removal of one gamma-subunit to give alpha beta gamma results in a Vmax. 5.2 +/- 1% of that for maximal activity. Frog alpha beta gamma 2 has a Vmax. 10.8 +/- 2%, and alpha beta gamma has a Vmax. 50 +/- 18%, of the Vmax. of alpha beta. These data suggest that a single gamma-subunit can inhibit the catalytic activity of active sites on both alpha- and beta-subunits in bovine, but not in frog, rod-outer-segment PDE.

scholarly journals Ouabain binding to phospholipid-dependent adenosine triphosphatase

1978 ◽  
Vol 169 (2) ◽  
pp. 313-320 ◽  
Author(s):  
S L Goodman ◽  
K P Wheeler
Keyword(s):  
Atpase Activity ◽  
Protein Complex ◽  
Adenosine Triphosphatase ◽  
Rabbit Kidney ◽  
Ouabain Binding ◽  
Equilibrium Conditions ◽  
Enzyme Preparations

The role of phospholipid in the binding of ouabain to the (Na+ + K+)-dependent adenosine triphosphatase was studied. Enzyme preparations obtained from rabbit kidney were treated with Lubrol WX to remove the phospholipid component essential for ATPase activity. Reconstituted enzyme samples were prepared by the addition of phosphatidylserine and sedimentation of an enzymically active lipid-protein complex. The binding of ouabain to both kinds of preparations was measured under equilibrium conditions with the use of 3H-labelled ouabain and initial ouabain concentrations in the range 0.01-1 micrometer. The main findings were: (i) (Mg2+ + Pi) promoted binding of significant quantities of ouabain only to the reconstituted enzyme; (ii) the absence of added Na+, (Mg2+ + ATP) similarly promoted binding only to the reconstituted samples; (iii) the addition of Na+ in the presence of (Mg2+ + ATP) increased the amount of ouabain bound to the reconstituted enzyme when the ouabain concentration was below about 0.1 micrometer, but it had no effect when the ouabain concentration was about 1 micrometer; (iv) (Mg2+ + ATP) induced ouabain binding to the depleted enzyme only when Na+ was also added; (v) the amount of ouabain bound to both depleted and reconstituted enzymes was the same in the presence of (Mg2+ + ATP + Na+); (vi) the reconstituted enzyme appeared to have a greater affinity for Na+ than did the depleted enzyme.

scholarly journals A new appraisal of the endoglucanases of the fungus Trichoderma reesei

1985 ◽  
Vol 231 (1) ◽  
pp. 75-81 ◽  
Author(s):  
M L Niku-Paavola ◽  
A Lappalainen ◽  
T M Enari ◽  
M Nummi
Keyword(s):  
Trichoderma Reesei ◽  
Specific Activity ◽  
Relative Proportion ◽  
Culture Liquid ◽  
Enzymic Activity ◽  
Soluble Form ◽  
Immunological Methods ◽  
A Value ◽  
And Storage

The properties and enzymic activity of endoglucanases (EC 3.2.1.4) of the fungus Trichoderma reesei were studied by means of immunological methods and by using polyglycosidic substrates. Endoglucanases exist in the culture liquid as a series of immunologically related components. The most active endoglucanase component has an Mr of 43 000 and pI value of 4.0. The most abundant components have a value of pI about 5.0, an Mr of 56 000-67 000 and specific activity only one-fifth of that of the pI-4.0 component. During purification and storage the endoglucanases are spontaneously modified; the relative proportion of components having greater Mr values, more alkaline pI values and lower specific activities is increased. The hexose content of the endoglucanase components is 2-7%. Endoglucanases hydrolyse soluble β-1,4 glycans. The enzymes described here differ from endoglucanase preparations described previously in not showing activity towards insoluble substrates. The role of endoglucanases in wood hydrolysis is consequently limited to the stage where wood constituents are already in soluble form.

scholarly journals Mitochondrial adenosine triphosphatase of the fission yeast, Schizosaccharomyces pombe 972h-. Changes in activity and inhibitor-sensitivity in response to catabolite repression

1976 ◽  
Vol 160 (2) ◽  
pp. 335-342 ◽  
Author(s):  
D Lloyd ◽  
S W Edwards
Keyword(s):  
Atpase Activity ◽  
Schizosaccharomyces Pombe ◽  
Adenosine Triphosphatase ◽  
Specific Activity ◽  
Early Stage ◽  
Gel Filtration ◽  
Growth Cycle ◽  
Mitochondrial Atpase ◽  
Submitochondrial Particles ◽  
Fourfold Increase

1. The specific activity of mitochondrial ATPase (adenosine triphosphatase) in extracts of Schizosaccharomyces pombe decreased 2.5-fold as the glucose concentration in the growth medium decreased from 50mM to 15mM. 2. During the late exponential phase of growth, ATPase activity doubled. 3. Sensitivity to oligomycin and Dio-9 as measured by values for I50(mug of inhibitor/mg of protein giving 50% inhibition) at pH 6.8 increased sixfold and ninefold respectively during the initial decrease in ATPase activity, and this degree of sensitivity was maintained for the remainder of the growth cycle. 4. Increased sensitivity to NN′-dicyclohexylcarbodi-imide, triethyltin and venturicidin was also observed during the early stage of glucose de-repression. 5. Smaller increases in sensitivity to efrapeptin, aurovertin, 7-chloro-4-nitrobenzo-2-oxa-1,3-diaz-le, quercetin and spegazzinine also occurred. 6. The ATPase of glycerol-grown cells was less sensitive to inhibitors than that of glucose-repressed cells; change in values for I50 were not so marked during the growth cycle of cells growing with glycerol. 7. When submitochondrial particles from glycerol-grown cells were tested by passage through Sephadex G-50, a fourfold increase in activity was accompanied by increased inhibitor resistance. 8. Gel filtration of submitochondrial particles from glucose-de-repressed cells gave similar results, whereas loss of ATPase occurred in submitochondrial particles from glucose-repressed cells. 9. It is proposed that alterations in sensitivity to inhibitors at different stages of glucose derepression may be partly controlled by a naturally occuring inhibitor of ATPase. 10. The inhibitors tested may be classififed into two groups on the basis of alterations of sensitivity of the ATPase during physiological modification: (a) oligomycin, Dio-9, NN′-dicyclohexylcarbodi-imide, venturicidin and triethyltin, and (b) efrapeptin, aurovertin, 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole, quercetin and spegazzinine.

Role of Na+/K+-stimulated adenosine triphosphatase in the action of dopaminergic-D2 receptors of the liver in rats

1987 ◽  
Vol 7 (11) ◽  
pp. 839-842 ◽  
Author(s):  
Abdulrahim Abu-Jayyab ◽  
Ahmed Mahgoub
Keyword(s):  
Atpase Activity ◽  
Receptor Antagonist ◽  
Dopamine Receptor ◽  
Inhibitory Activity ◽  
Receptor Agonist ◽  
Adenosine Triphosphatase ◽  
D2 Receptors ◽  
Dopamine Receptor Agonist ◽  
Dopamine Receptor Antagonist

The dopamine receptor agonist, bromocriptine, in a dose of 10 mg/kg i.p. for 14 days, in rats caused a significant increase in liver Na+/K+-ATPase activity, whereas sulpiride, a dopamine receptor antagonist, in a dose of 10 mg/kg, i.p. for 14 days, in rats, caused a significant decrease in liver Na+/K+-ATPase activity. Injection of bromocriptine and sulpiride simultaneously in a group of rats, under the same conditions and using the same doses caused a complete block of both stimulatory activity of bromocriptine and inhibitory activity of sulpiride on liver Na+/K+-ATPase activity. It is suggested that Na+/K+-ATPase may have a role in the action of dopaminergic-D2 receptors.

UTERINE MITOSIS, ALKALINE PHOSPHATASE AND ADENOSINE TRIPHOSPHATASE DURING DEVELOPMENT AND REGRESSION OF DECIDUOMATA IN PSEUDOPREGNANT MICE

1969 ◽  
Vol 44 (1) ◽  
pp. 91-NP ◽  
Author(s):  
KATHLEEN HALL
Keyword(s):  
Alkaline Phosphatase ◽  
Atpase Activity ◽  
Vascular Endothelium ◽  
Stromal Cells ◽  
Adenosine Triphosphatase ◽  
Glandular Epithelium ◽  
Histochemical Localization ◽  
Vessel Walls ◽  
The Right

SUMMARY Incidence and patterns of mitoses and histochemical localization of alkaline phosphatase (APase) and adenosine triphosphatase (ATPase) were studied from days 5 to 20 in pseudopregnant mice in which deciduomata had been induced in the left uterine horns by intrauterine injection of oil on day 4, the right horns serving as controls. In stromal cells, mitoses were very numerous throughout the endometrium of the left (but not the right) horn on days 5 and 6, in the basal, non-decidualized stroma until day 8 or day 9, and were not seen in stromal cells of either horn thereafter. In glandular epithelium, mitoses were absent from days 5 to 10, and numerous from day 11 or 12 until at least day 17 in both horns. Mitoses were present in capillaries within developing deciduomata on days 5 and 6, then seldom seen until day 12 and during the next 3 days were numerous in endothelial and pericapillary cells in the mesometrial quadrant of the left horn and around glands in both horns. The deciduoma cells reacted strongly with AP and ATP substrates from days 5 to 10, after which the intensity of the reaction weakened and had usually disappeared by day 13. ATPase activity disappeared from vascular endothelium within the deciduoma a few hours after APase had appeared within the deciduoma cells on day 5. It reappeared in the vessel walls on day 9 and thereafter was usually present until the deciduoma was shed. In the basal, non-decidualized stroma, APase was absent until about day 10, then appeared in the stroma cells nearest to the myometrium, extending gradually into the densely packed cells nearest to the regressing deciduoma. The possible role of this enzyme in reparative growth of the endometrium after regression of the deciduoma is discussed.

scholarly journals Regulation of Dopamine Uptake by Vasoactive Peptides in the Kidney

Scientifica ◽  
2016 ◽  
Vol 2016 ◽  
pp. 1-12
Author(s):  
N. L. Rukavina Mikusic ◽  
N. M. Kouyoumdzian ◽  
E. Rouvier ◽  
M. M. Gironacci ◽  
J. E. Toblli ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Inhibitory Effect ◽  
Renal Tissue ◽  
Dopamine Uptake ◽  
Tubular Cells ◽  
Vasoactive Peptides ◽  
Renal Dopamine ◽  
Atpase Inhibition ◽  
Sodium Handling

Considering the key role of renal dopamine in tubular sodium handling, we hypothesized that c-type natriuretic peptide (CNP) and Ang-(1-7) may regulate renal dopamine availability in tubular cells, contributing to Na+, K+-ATPase inhibition. Present results show that CNP did not affect either3H-dopamine uptake in renal tissue or Na+, K+-ATPase activity; meanwhile, Ang-(1-7) was able to increase3H-dopamine uptake and decreased Na+, K+-ATPase activity in renal cortex. Ang-(1-7) and dopamine together decreased further Na+, K+-ATPase activity showing an additive effect on the sodium pump. In addition, hydrocortisone reversed Ang-(1-7)-dopamine overinhibition on the enzyme, suggesting that this inhibition is closely related to Ang-(1-7) stimulation on renal dopamine uptake. Both anantin and cANP (4-23-amide) did not modify CNP effects on3H-dopamine uptake by tubular cells. The Mas receptor antagonist, A-779, blocked the increase elicited by Ang-(1-7) on3H-dopamine uptake. The stimulatory uptake induced by Ang-(1-7) was even more pronounced in the presence of losartan, suggesting an inhibitory effect of Ang-(1-7) on AT1 receptors on3H-dopamine uptake. By increasing dopamine bioavailability in tubular cells, Ang-(1-7) enhances Na+, K+-ATPase activity inhibition, contributing to its natriuretic and diuretic effects.

scholarly journals The adenosine triphosphatase and calcium ion-transporting activities of the sarcoplasmic reticulum of developing muscle

1969 ◽  
Vol 114 (2) ◽  
pp. 161-170 ◽  
Author(s):  
D. L. Holland ◽  
S V Perry
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Atpase Activity ◽  
Adenosine Triphosphatase ◽  
Specific Activity ◽  
Sucrose Density Gradient ◽  
Maximum Activity ◽  
Longissimus Dorsi ◽  
Longissimus Dorsi Muscle ◽  
Dorsi Muscle ◽  
Coupling System

1. The ATPase (adenosine triphosphatase) specific activity and the total nitrogen content of the myofibrillar fraction per g. wet weight of rabbit longissimus dorsi muscle increased steadily during the late foetal stages and the first few weeks after birth. 2. The ATPase specific activity of the sarcoplasmic-reticular fraction isolated by a sucrose-density-gradient procedure rose to a sharp peak 8–10 days after birth and then declined to the adult value, which was about 25% of the maximum. 3. The peak in ATPase activity was a feature of the sarcoplasmic reticulum isolated from muscle, and the time at which it occurred in relation to birth was related to the degree of development and the activity pattern of the muscle. 4. The peak in ATPase activity of the sarcoplasmic reticulum occurred at an earlier age if newborn animals were made to exercise earlier than was normal. 5. The ‘extra’ ATPase associated with the sarcoplasmic reticulum and the ability to concentrate Ca2+ increased in a similar manner over the period of development studied. 6. It is postulated that the Ca2+-transport system of the sarcoplasmic reticulum consists of two components, namely the ATPase and the system coupling this enzyme to Ca2+ transport. During development the ATPase develops first and has almost reached maximum activity in the longissimus dorsi muscle of the rabbit after 8–10 days. Subsequently the activity of the coupling system rises rapidly, leading to an increase in the capacity and efficiency of Ca2+ transport.

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