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Glucose-6-phosphate dehydrogenase. Purification and partial characterization.

M I Kanji, M L Toews, W R Carper

    The Journal of Biological Chemistry
    |April 25, 1976
    PubMed
    Summary

    Pig liver glucose-6-phosphate dehydrogenase was purified, revealing an active dimer and inactive monomer. This soluble cytoplasmic enzyme shows high specificity for NADP+ and glucose 6-phosphate.

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    Area of Science:

    • Biochemistry
    • Enzymology

    Background:

    • Glucose-6-phosphate dehydrogenase (G6PD) is a critical enzyme in cellular metabolism.
    • Understanding G6PD's properties is essential for metabolic pathway research.

    Purpose of the Study:

    • To purify and characterize glucose-6-phosphate dehydrogenase from pig liver.
    • To determine the enzyme's molecular weight, optimal activity conditions, substrate specificity, and cellular localization.

    Main Methods:

    • Enzyme purification using biochemical techniques (1000-fold purification).
    • Determination of molecular weight via ultracentrifugation or gel filtration.
    • Assays to determine optimal pH, ionic strength, substrate specificity, and kinetic parameters (Km).
    • Cellular fractionation to ascertain enzyme localization.

    Main Results:

    • Purified G6PD exists as an active dimer (133,000 MW) and an inactive monomer (67,500 MW).
    • Optimal activity occurs at pH 8.5 and ionic strength of 0.1–0.5 M.
    • The enzyme exhibits high specificity for NADP+ and glucose 6-phosphate, with Km values of 3.6 µM and 5.4 µM, respectively.
    • G6PD is predominantly found in the soluble cytoplasmic fraction.

    Conclusions:

    • The study characterizes pig liver G6PD, detailing its oligomeric states and enzymatic properties.
    • Findings provide insights into G6PD's role and regulation within the cellular cytoplasm.
    • This work contributes to the biochemical understanding of G6PD function.

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