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Related Experiment Videos

Yeast inorganic pyrophosphatase substrate recognition.

W B Knight, S J Ting, S Chuang

    Archives of Biochemistry and Biophysics
    |November 1, 1983
    PubMed
    Summary

    Yeast inorganic pyrophosphatase acts on bidentate pyrophosphate complexes, with activity influenced by metal-ligand composition. Metal-water ligands play a role in productive binding and hydrolysis rates, with varying effects under Mg2+ and Zn2+ activation.

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    Biophysical journal·2009

    Area of Science:

    • Biochemistry
    • Enzymology
    • Inorganic Chemistry

    Background:

    • Inorganic pyrophosphatase (IPPase) is a crucial enzyme in cellular metabolism, catalyzing pyrophosphate hydrolysis.
    • The substrate specificity and catalytic mechanisms of IPPases are influenced by the structure and coordination of pyrophosphate complexes.

    Purpose of the Study:

    • To investigate the substrate activity of various metal-dipyrophosphate complexes with yeast inorganic pyrophosphatase.
    • To elucidate the role of metal-ligand composition, particularly metal-aqua ligands, in enzyme binding and catalysis.
    • To compare the activity of Mg2+-activated and Zn2+-activated enzyme towards different pyrophosphate complexes.

    Main Methods:

    • Synthesis and characterization of various metal-dipyrophosphate complexes, including cobalt and chromium complexes with varying ligands.

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  • Enzymatic assays using yeast inorganic pyrophosphatase to determine kinetic parameters (Km, kcat) for substrate hydrolysis.
  • Investigation of enzyme activity under different metal ion activations (Mg2+ and Zn2+).

    • Main Results:

    • Monodentate Co(NH3)5PPi is not a substrate, while P1,P2-bidentate Co(NH3)4PPi is hydrolyzed at 7.5 min-1.
    • For P1,P2-bidentate Cr-PPi complexes, increasing ammonia substitution for water ligands decreased binding affinity and turnover number.
    • Mg2+-activated enzyme did not hydrolyze beta, gamma-bidentate or alpha, beta, gamma-tridentate Co-PPPi complexes, but Zn2+-activated enzyme hydrolyzed the tridentate complex at 0.17 min-1.

    Conclusions:

    • Yeast inorganic pyrophosphatase exhibits specificity towards bidentate pyrophosphate complexes, with activity modulated by metal coordination.
    • Metal-aqua ligands influence substrate binding and catalytic efficiency, with ammonia substitution reducing activity.
    • The enzyme's metal ion cofactor (Mg2+ vs. Zn2+) significantly impacts its substrate specificity and hydrolysis capability for different pyrophosphate coordination modes.