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Identification of ATP diphosphohydrolase activity in human term placenta using a novel assay for AMP

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Abstract

Human term placenta contains an ATP diphosphohydrolase activity which hydrolyses ATP to ADP and inorganic phosphate and ADP to AMP and a second mole of inorganic phosphate. The activity has a pH optimum between 8.0 and 8.5. Magnesium or calcium ions are required for maximum activity. Other nucleoside phosphates, p-nitrophenyl phosphate or sodium pyrophosphate, are not hydrolysed. The activity is not due to ATPases, or to myokinase, as determined by the use of inhibitors. NaF and NaN3 were found to inhibit strongly the activity thus identifying it as an ATP diphosphohydrolase.

A sensitive enzymatic assay for measurement of AMP, one of the products of the reaction, was established, based on the strong inhibition of muscle fructose 1,6-biphosphatase by AMP. The range of the assay was 0.05–0.8 µM AMP. ATP diphosphohydrolase was found to have a rate of AMP production from ADP twice the rate from ATP. Under the same conditions, the assay for Pi release, on the other hand, gave velocities similar to each other for the two substrates.

The activity appears to be identical to the ADP-hydrolysing activity in placenta reported by others.

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Abbreviations

Ap5A:

P1

P5-di(adenosine-5′):

Pentaphosphate

ATP-DPH:

ATP Diphosphohydrolase

DCCD:

N,N′ Dicyclohexycarbodiimide

Fru-P2ase:

Fructose 1,6-biphosphatase

SDS:

Sodium Dodecyl Sulfate

TLC:

Thin Layer Chromatography

References

  1. Kalckar HM: Adenylpyrophosphatase and myokinase. J Biol Chem 153: 355–367, 1944

    Google Scholar 

  2. Knowles AF, Isler RE, Reece JF: The common occurrence of ATP diphosphohydrolase in mammalian plasma membranes. Biochim Biophys Acta 731: 88–96, 1983

    Google Scholar 

  3. LeBel D, Poirier GG, Phaneuf S, St-Jean P, Laliberte JF, Beaudoin AR: Characterization and purification of a calcium-sensitive ATP diphosphohydrolase from pig pancreas. J Biol Chem 255: 1227–1233, 1980

    Google Scholar 

  4. Laliberte JF, St-Jean P, Beaudoin AR: Kinetic effects of Ca2+ and Mg2+ on ATP hydrolysis by the purified ATP diphosphohydrolase. J Biol Chem 257:3869–3874, 1982

    Google Scholar 

  5. Laliberte JF, Beaudoin AR: Sequential hydrolysis of the y and β-phosphate groups of ATP by the ATP diphosphohydrolase from pig pancreas. Biochim Biophys Acta 742: 9–15, 1983

    Google Scholar 

  6. Miura Y, Hirota K, Arai Y, Yagi K: Purification and partial characterization of adenosine diphosphatase activity in bovine aorta microsomes. Thromb Res 46: 685–695, 1987

    Google Scholar 

  7. Yagi K, Arai Y, Kato N, Hirota K, Miura Y: Purification of ATP diphosphohydrolase from bovine aorta microsomes. Eur J Biochem 180: 509–513, 1989

    Google Scholar 

  8. Rosenberg MD, Buegel J, Strobel RS: Purification and characterization of nucleotide diphosphohydrolase from oviductal and seminal secretions. J Cell Biol 107:178 Abstr, 1988

    Google Scholar 

  9. Hutton RA, Chow FPR, Craft IL, Dandona P: Inhibitors of platelet aggregation in the fetoplacental unit and myometrium with particular reference to the ADP-degrading property of placenta. Placenta 1: 125–130, 1980

    Google Scholar 

  10. Hutton RA, Dandona P, Cow FPR, Craft IL: Inhibition of platelet aggregation by placental extracts. Thromb Res 17: 465–471, 1980

    Google Scholar 

  11. Black WJ, Van Tol A, Fernando J, Horecker BL: Isolation of a higly active fructose diphosphatase from rabbit muscle: Its subunit structure and activation by monovalent cations. Arch Biochem Biophys 151: 576–590, 1972

    Google Scholar 

  12. Pontremoli S, Traniello S, Luppis B, Wood WA: Fructose diphosphatase from rabbit liver. 1. Purification and properties. J Biol Chem 240: 3459–3463, 1965

    Google Scholar 

  13. Annamalai AE, Tsolas O, Horecker BL: fructose 1,6-bisphosphatase from chicken breast muscle. Arch Biochem Biophys 183: 48–56, 1977

    Google Scholar 

  14. Horecker BL, Kornberg A: The extinction coefficients of the reduced band of pyridine nucleotides. J Biol Chem 175: 385–389, 1948

    Google Scholar 

  15. Fiske CH, Subbarow Y: The colorimetric determination of phosphorus. J Biol Chem 66: 375–400, 1925

    CAS  Google Scholar 

  16. Randerath K, Struck H: Dunnschichtchromatographische Trennung von Nucleinsaure-derivaten an Celluloseschichten. J Chromatog 6: 365–367, 1961

    Google Scholar 

  17. Randerath K: A comparison between thin-layer chromatography and paper chromatography of nucleic acid derivatives. Biochem Biophys Res Commun 6: 452–457, 1962

    Google Scholar 

  18. Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254, 1976

    Article  CAS  PubMed  Google Scholar 

  19. Lowry OH, Rosebrough NJ, Farr LA, Randall RJ: Protein measurement with the Folin phenol reagent. J Biol Chem 193: 265–275, 1951

    CAS  PubMed  Google Scholar 

  20. Macguire MH, Krishnakantha TP, Aronson DM: Human placental 5′-nucleotidase: Purification and properties. Placenta 5: 21–40, 1984

    Google Scholar 

  21. Cori O, Traverso-Cori A, Montserrat T, Chaimovich H: Substrate specificity and inhibition studies on potato apyrase: Biochem Z 342: 345–358, 1965

    Google Scholar 

  22. Jorgensen PL: Purification and characterization of (Na+ + K+)-ATPase. IV. Estimation of the purity and of the molecular weight of polypeptide content per enzyme unit in preparations from the outer medulla of rabbit kidney. Biochem Biophys Acta 356: 53–67, 1974

    Google Scholar 

  23. Gottschalk A, Bhargava AS: Neuraminidases. In: the Enzymes, 3rd. edn. (P.D. Boyer, ed.), vol. V: pp. 321–342, Academic Press, New York, 1971

  24. Beechy RB, Roberton AM, Holloway CT, Knight IG: The properties of dicyclohexylcarbodiimide as an inhibitor of oxidative phosphorylation. Biochemistry 6: 3867–3879, 1967

    Google Scholar 

  25. Ghosh NK, Fishman WH: On the mechanism of inhibition of intestinal alkaline phosphatase by L-phenylalanine. J Biol Chem 241: 2516–2522, 1966

    Google Scholar 

  26. Meyerhof O: The origin of the reaction of Harden and Young in cell-free alcoholic fermentation. J Biol Chem 157: 105–119, 1945

    Google Scholar 

  27. Lienhard GE, Secemscki II: P1,P5-Di(adenosine-5′)penta-phosphate, a potent multisubstrate inhibitor of adenylate kinase. J Biol Chem 248: 1121–1123, 1973

    Google Scholar 

  28. Eggleston LV, Hems R: Separation of adenosine phosphates by paper chromatography and the equilibrium constant of the myokinase system. Biochem J 52:156–160, 1952

    Google Scholar 

  29. Montague DJ, Peters TJ, Baum H: Comparison of effects of inhibitors on adenosine triphosphatase and adenosine diphosphatase activities in rat-liver mitochondria. Eur J Biochem 139:529–534, 1984

    Google Scholar 

  30. Luthjie J, Schomburg A, Ogilvie A: Demonstration of a novel ecto-enzyme on human erythrocytes, capable of degrading ADP and of inhibiting ADP-induced platelet aggregation. Eur J Biochem 175: 285–289, 1988

    Google Scholar 

  31. Gaardner A, Jonsen J, Laland S, Hellem A, Owren PA: Adenosine diphosphate in red cells as a factor in the adhesiveness of human blood platelets. Nature 192: 531–532, 1961

    Google Scholar 

  32. Born GVR: Aggregation of blood platelets by adenosine diphosphate and its reversal. Nature 194: 927–929, 1962

    Google Scholar 

  33. Smith JJB, Cornish RA, Wilkes J: Properties of a calcium-dependent apyrase in the saliva of the blood-feeding bug, Rhodnius prolixus. Experientia 36: 898–900, 1980

    Google Scholar 

  34. Sarkis JIF, Guimaraes JA, Ribeiro JMC: Salivary apyrase of Rhodnius prolixus. Biochem J 233: 885–891, 1986

    Google Scholar 

  35. Beaudoin AR, Vachereau A, Grondin G, St Jean P, Rosenberg MD, Strobel R: Microvesicular secretion, a mode of cell secretion associated with the presence of an ATP-diphosphohydrolase. FEBS Lett 203: 1–2, 1986

    Google Scholar 

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  1. Laboratory of Biological Chemistry, University of Ioannina Medical School, P. O. Box 1186, GR-451 10, Ioannina, Greece

    Thomais Papamarcaki & Orestes Tsolas

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  1. Thomais Papamarcaki
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  2. Orestes Tsolas
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Papamarcaki, T., Tsolas, O. Identification of ATP diphosphohydrolase activity in human term placenta using a novel assay for AMP. Mol Cell Biochem 97, 1–8 (1990). https://doi.org/10.1007/BF00231696

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  • DOI: https://doi.org/10.1007/BF00231696

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