Characterization of the pyrogallol-phloroglucinol isomerase of Eubacterium oxidoreducens
Cell extracts of Eubacterium oxidoreducens, in the presence of dimethyl sulfoxide, catalyzed the conversion of pyrogallol to phloroglucinol with methyl sulfide as a product. The isomerization reaction also proceeded when 1,2,3,5-benzenetetrol was present rather than dimethyl sulfoxide. An assay to quantitate this activity was developed. The assay followed the disappearance of 1,2,4-benzenetriol as determined colorimetrically after incubation with sodium molybdate at neutral pH. The products of this reaction were resorcinol and 2,6-dihydroxyquinone. The enzyme(s) catalyzing this reaction was purified fivefold from cells grown on gallate plus H/sub 2/. The purification procedure involved treatment with 40% acetone, precipitation with ammonium sulfate, DEAE-cellulose chromatography, concentration by ultrafiltration (molecular weight cutoff, > 100,000), and hydroxylapatite chromatography. This preparation had a specific activity of 14.7 ..mu..mol/min per mg of protein and a pH optimum of about 7.3. It was strongly inhibited by p-chloromercuribenzoate. The mechanism of the reaction involved oxidation of the pyrogallol followed by introduction of water. The benzenetetrol intermediate was then reduced and dehydrated to phloroglucinol.
- Research Organization:
- Univ. of Illinois, Urbana (USA)
- DOE Contract Number:
- AC02-81ER10874
- OSTI ID:
- 6769435
- Journal Information:
- J. Bacteriol.; (United States), Vol. 170:6
- Country of Publication:
- United States
- Language:
- English
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