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Cytochrome P-450-dependent fragmentation of DNA in reconstituted membranes

H Luthman, M Ingelman-Sundberg

    Acta Pharmacologica Et Toxicologica
    |January 1, 1985
    PubMed

    Insights

    Rabbit liver microsomes containing cytochromes P-450 degrade plasmid DNA via NADPH-dependent hydroxyl radical generation. This process, inhibited by scavengers, involves an iron-catalyzed Haber-Weiss reaction.

    Area of Science:

    • Biochemistry
    • Molecular Biology
    • Enzymology

    Background:

    • Microsomal enzymes, including cytochromes P-450 (CYP450), play crucial roles in cellular metabolism.
    • Reactive oxygen species (ROS) generation is associated with various biological processes and cellular damage.
    • The interaction between CYP450 enzymes and nucleic acids is an area of ongoing research.

    Purpose of the Study:

    • To investigate the potential of rabbit liver microsomes and associated enzymes to degrade plasmid DNA.
    • To elucidate the mechanism of DNA degradation, specifically identifying the reactive species involved.
    • To determine the role of specific cytochromes and cofactors in this process.

    Main Methods:

    • Incubation of plasmid DNA with rabbit liver microsomes containing various forms of cytochromes P-450 and NADPH-cytochrome P-450 reductase.
    • Assessing DNA degradation in the presence and absence of NADPH.
    • Replacing cytochrome P-450 with cytochrome b5 to evaluate its role.
    • Utilizing hydroxyl radical scavengers, superoxide dismutase, and catalase to inhibit the reaction.

    Main Results:

    • Membrane vesicles containing cytochromes P-450 and NADPH-cytochrome P-450 reductase degraded plasmid DNA in an NADPH-dependent manner.
    • Replacement of cytochrome P-450 with cytochrome b5 resulted in negligible DNA disintegration.
    • Complete inhibition of DNA degradation was observed with hydroxyl radical scavengers, superoxide dismutase, and catalase.
    • These findings suggest the generation of hydroxyl radicals via an iron-catalyzed Haber-Weiss reaction.

    Conclusions:

    • Rabbit liver microsomal cytochromes P-450, in conjunction with NADPH-cytochrome P-450 reductase, can induce DNA degradation.
    • Hydroxyl radicals, generated through an iron-catalyzed Haber-Weiss reaction, are the primary mediators of this DNA damage.
    • Cytochrome b5 does not significantly contribute to the DNA degradation process mediated by CYP450.

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