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Dinucleosidetriphosphatase from rat brain.

M J Costas, J M Montero, J C Cameselle

    The International Journal of Biochemistry
    |January 1, 1984
    PubMed
    Summary
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    Researchers purified rat brain adenylohydrolase (dinucleosidetriphosphatase) which hydrolyzes diadenosine and diguanosine triphosphates. This enzyme requires specific divalent cations for activity and has an optimal pH of 7.5.

    Area of Science:

    • Biochemistry
    • Enzymology
    • Molecular Biology

    Background:

    • Diadenosine triphosphates (Ap3A) are signaling molecules involved in various cellular processes.
    • Adenylohydrolase (dinucleosidetriphosphatase, EC 3.6.1.29) plays a role in regulating Ap3A levels.
    • Understanding the kinetic and catalytic properties of this enzyme is crucial for elucidating its biological function.

    Purpose of the Study:

    • To purify and characterize adenylohydrolase (dinucleosidetriphosphatase) from rat brain.
    • To determine the substrate specificity and kinetic parameters of the purified enzyme.
    • To investigate the effects of metal ions and pH on enzyme activity.

    Main Methods:

    • 1000-fold purification of adenylohydrolase from rat brain.
    • Enzyme activity assays using various diadenosine and diguanosine polyphosphates.

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  • Determination of kinetic parameters (Km, Vmax) and inhibition constants (Ki).
  • Analysis of substrate specificity, pH optimum, molecular mass, and isoelectric point.

    • Main Results:

    • The purified enzyme efficiently hydrolyzed diadenosine triphosphate (Ap3A) and diguanosine triphosphate (Gp3G) to their respective nucleoside di- and monophosphates.
    • Km values were 14 microM for Ap3A and 40 microM for Gp3G, with relative Vmax of 100 and 40, respectively.
    • The enzyme required Mg(II), Mn(II), or Ca(II) for activity, with Zn(II) acting as a competitive inhibitor (Ki = 5 microM).
    • Optimum pH was approximately 7.5, molecular mass was 34 kDa, and isoelectric point was 5.5.

    Conclusions:

    • Rat brain adenylohydrolase is a specific dinucleosidetriphosphatase with distinct substrate preferences.
    • The enzyme's activity is modulated by divalent cations and pH, providing insights into its physiological regulation.
    • Characterization of this enzyme contributes to understanding the metabolism of Ap3A and related signaling molecules.