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p-cymene pathway in Pseudomonas putida: initial reactions

J J DeFrank, D W Ribbons

    Journal of Bacteriology
    |March 1, 1977
    PubMed
    Summary

    Pseudomonas putida PL metabolizes p-cymene via a pathway involving a dihydrodiol intermediate, not 3-hydroxy-p-cumate. This intermediate is crucial for ring cleavage and bacterial growth on p-cymene.

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    Area of Science:

    • Microbial Metabolism
    • Biochemistry
    • Environmental Microbiology

    Background:

    • The p-cymene degradation pathway in Pseudomonas putida is essential for breaking down aromatic compounds.
    • Previous understanding suggested 3-hydroxy-p-cumate as a key intermediate in this pathway.

    Purpose of the Study:

    • To reinvestigate and clarify the initial steps of the p-cymene pathway in Pseudomonas putida PL.
    • To identify the correct intermediate preceding ring cleavage in p-cymene metabolism.

    Main Methods:

    • Utilized a mutant strain (P. putida PL-pT-11/43) unable to grow on p-cymene to identify accumulated metabolites.
    • Employed enzyme assays with nicotinamide adenine dinucleotide-dependent dehydrogenase.
    • Conducted mass spectral analysis of dihydrodiol intermediates using 18O2-enriched atmospheres.

    Main Results:

    • Identified 2,3-dihydroxy-4-isopropylcyclohexa-4,6-dienoate as a dihydrodiol intermediate, not 3-hydroxy-p-cumate.
    • Demonstrated the enzymatic oxidation of this dihydrodiol by a specific dehydrogenase present in wild-type and revertant strains.
    • Confirmed that 3-hydroxy-p-cumate does not support growth or oxidation, suggesting it's not a physiological intermediate.

    Conclusions:

    • The p-cymene pathway proceeds via a dihydrodiol intermediate, which is enzymatically oxidized before ring cleavage.
    • This dihydrodiol is the direct precursor for the ring cleavage enzyme, revising the previously accepted pathway.
    • The study provides strong evidence for the physiological role of the dihydrodiol intermediate in p-cymene biodegradation.

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