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Isolation and Functional Analysis of Mitochondria from Cultured Cells and Mouse Tissue
09:27

Isolation and Functional Analysis of Mitochondria from Cultured Cells and Mouse Tissue

Published on: March 23, 2015

Phosphorylation of adenosine monophosphate in the mitochondrial matrix

H A Krebs, D Wiggins

    The Biochemical Journal
    |July 15, 1978
    PubMed

    Insights

    Investigating GTP origin in rat hepatocytes revealed that nucleoside diphosphokinase significantly contributes to GTP supply for AMP phosphorylation within the mitochondrial matrix.

    Area of Science:

    • Biochemistry
    • Mitochondrial Metabolism

    Background:

    • Adenosine monophosphate (AMP) is formed in the mitochondrial matrix during short-chain fatty acid metabolism via butyryl-CoA ligase.
    • GTP supply is crucial for the reconversion of AMP to ADP by GTP-AMP transphosphorylase.

    Purpose of the Study:

    • To investigate the origin of guanosine triphosphate (GTP) utilized for adenosine monophosphate (AMP) phosphorylation in the mitochondrial matrix.
    • To determine the relative contributions of succinic thiokinase and nucleoside diphosphokinase to GTP synthesis.

    Main Methods:

    • Quantified matrix AMP formation rate by measuring ketone body production in rat hepatocytes.
    • Calculated succinic thiokinase reaction rate using tricarboxylic acid cycle turnover, derived from oxygen consumption and ketone body formation.
    • Assessed GTP supply rates from both succinic thiokinase and nucleoside diphosphokinase.

    Main Results:

    • Matrix AMP formation rate was calculated from ketone body formation rates.
    • Nucleoside diphosphokinase was found to contribute up to 80% to the GTP supply under tested conditions.
    • The rate of GTP-AMP transphosphorylase activity is limited by GTP availability.

    Conclusions:

    • Nucleoside diphosphokinase plays a major role in mitochondrial GTP synthesis.
    • Understanding GTP origin is key to comprehending AMP phosphorylation regulation in hepatocytes.
    • Metabolic flux analysis provides insights into mitochondrial enzyme contributions to nucleotide pools.

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