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Asparagine utilization in Escherichia coli.

R C Willis, C A Woolfolk

    Journal of Bacteriology
    |April 1, 1974
    PubMed
    Summary
    This summary is machine-generated.

    Escherichia coli K-12 auxotrophs degrade excess asparagine supplements instead of conserving them for protein synthesis. Supplement conservation occurs when asparagine degradation is blocked by mutation or the analogue DONV.

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    Proceedings of the National Academy of Sciences of the United States of America·1989

    Area of Science:

    • Microbiology
    • Molecular Biology
    • Biochemistry

    Background:

    • Asparagine is an essential amino acid for protein synthesis in Escherichia coli.
    • Asparagine-requiring auxotrophs rely on external supplements for growth.
    • Cytoplasmic asparaginase plays a role in asparagine metabolism.

    Purpose of the Study:

    • To investigate the conservation and degradation of asparagine supplements in Escherichia coli K-12.
    • To understand the role of cytoplasmic asparaginase in asparagine utilization.
    • To evaluate the effect of 5-diazo-4-oxo-l-norvaline (DONV) on asparagine metabolism.

    Main Methods:

    • Utilized asparagine-requiring auxotrophs of Escherichia coli K-12.
    • Assessed asparagine conservation under conditions of active cytoplasmic asparaginase.

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  • Investigated supplement conservation with inhibited asparagine degradation using DONV or mutation.
  • Determined asparagine requirements for protein synthesis in deficient strains.
  • Characterized cytoplasmic asparaginase kinetics (Km) and inhibition by DONV (Ki).

    • Main Results:

    • Auxotrophs with active cytoplasmic asparaginase do not conserve excess asparagine supplements; they are degraded.
    • Asparagine supplements are conserved when degradation is inhibited by DONV or mutation to asparaginase deficiency.
    • A strain deficient in cytoplasmic asparaginase required 260 mumol of asparagine per gram of cellular protein.
    • Cytoplasmic asparaginase (asparaginase I) is essential for growth when asparagine is the sole nitrogen source.
    • DONV competitively inhibits cytoplasmic asparaginase with an apparent Ki of 2 mM and causes time-dependent, irreversible inhibition in the absence of asparagine.

    Conclusions:

    • Cytoplasmic asparaginase actively degrades excess asparagine, preventing its accumulation for protein synthesis in Escherichia coli K-12.
    • Inhibition of asparaginase activity or deficiency is necessary for the conservation of asparagine supplements.
    • The characterized kinetic parameters and DONV inhibition provide insights into cytoplasmic asparaginase function and potential therapeutic targeting.