Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1996 Nov;178(21):6227–6232. doi: 10.1128/jb.178.21.6227-6232.1996

2-aminophenol 1,6-dioxygenase: a novel aromatic ring cleavage enzyme purified from Pseudomonas pseudoalcaligenes JS45.

U Lendenmann 1, J C Spain 1
PMCID: PMC178494  PMID: 8892823

Abstract

Most bacterial pathways for the degradation of aromatic compounds involve introduction of two hydroxyl groups either ortho or para to each other. Ring fission then occurs at the bond adjacent to one of the hydroxyl groups. In contrast, 2-aminophenol is cleaved to 2-aminomuconic acid semialdehyde in the nitrobenzene-degrading strain Pseudomonas pseudoalcaligenes JS45. To examine the relationship between this enzyme and other dioxygenases, 2-aminophenol 1,6-dioxygenase has been purified by ethanol precipitation, gel filtration, and ion exchange chromatography. The molecular mass determined by gel filtration was 140,000 Da. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed two subunits of 35,000 and 39,000 Da, which suggested an alpha2beta2 subunit structure. Studies with inhibitors indicated that ferrous iron was the sole cofactor. The Km values for 2-aminophenol and oxygen were 4.2 and 710 microM, respectively. The enzyme catalyzed the oxidation of catechol, 6-amino-m-cresol, 2-amino-m-cresol, and 2-amino-4-chlorophenol. 3-Hydroxyanthranilate, protocatechuate, gentisate, and 3- and 4-methylcatechol were not substrates. The substrate range and the subunit structure are unique among those of the known ring cleavage dioxygenases.

Full Text

The Full Text of this article is available as a PDF (213.7 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arciero D. M., Orville A. M., Lipscomb J. D. Protocatechuate 4,5-dioxygenase from Pseudomonas testosteroni. Methods Enzymol. 1990;188:89–95. doi: 10.1016/0076-6879(90)88017-5. [DOI] [PubMed] [Google Scholar]
  2. Cain R. B., Farr D. R. Metabolism of arylsulphonates by micro-organisms. Biochem J. 1968 Feb;106(4):859–877. doi: 10.1042/bj1060859. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. DAGLEY S., EVANS W. C., RIBBONS D. W. New pathways in the oxidative metabolism of aromatic compounds by microorganisms. Nature. 1960 Nov 12;188:560–566. doi: 10.1038/188560a0. [DOI] [PubMed] [Google Scholar]
  4. Furukawa K., Arimura N. Purification and properties of 2,3-dihydroxybiphenyl dioxygenase from polychlorinated biphenyl-degrading Pseudomonas pseudoalcaligenes and Pseudomonas aeruginosa carrying the cloned bphC gene. J Bacteriol. 1987 Feb;169(2):924–927. doi: 10.1128/jb.169.2.924-927.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ghosal D., You I. S., Gunsalus I. C. Nucleotide sequence and expression of gene nahH of plasmid NAH7 and homology with gene xylE of TOL pWWO. Gene. 1987;55(1):19–28. doi: 10.1016/0378-1119(87)90244-7. [DOI] [PubMed] [Google Scholar]
  6. Hallas L. E., Alexander M. Microbial transformation of nitroaromatic compounds in sewage effluent. Appl Environ Microbiol. 1983 Apr;45(4):1234–1241. doi: 10.1128/aem.45.4.1234-1241.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Harayama S., Kok M., Neidle E. L. Functional and evolutionary relationships among diverse oxygenases. Annu Rev Microbiol. 1992;46:565–601. doi: 10.1146/annurev.mi.46.100192.003025. [DOI] [PubMed] [Google Scholar]
  8. Harpel M. R., Lipscomb J. D. Gentisate 1,2-dioxygenase from Pseudomonas. Substrate coordination to active site Fe2+ and mechanism of turnover. J Biol Chem. 1990 Dec 25;265(36):22187–22196. [PubMed] [Google Scholar]
  9. Harpel M. R., Lipscomb J. D. Gentisate 1,2-dioxygenase from pseudomonas. Purification, characterization, and comparison of the enzymes from Pseudomonas testosteroni and Pseudomonas acidovorans. J Biol Chem. 1990 Apr 15;265(11):6301–6311. [PubMed] [Google Scholar]
  10. Hirata F., Nakazawa A., Nozaki M., Hayaishi O. Studies on metapyrocatechase. IV. Circular dichroism and optical rotatory dispersion. J Biol Chem. 1971 Oct 10;246(19):5882–5887. [PubMed] [Google Scholar]
  11. Kobayashi T., Ishida T., Horiike K., Takahara Y., Numao N., Nakazawa A., Nakazawa T., Nozaki M. Overexpression of Pseudomonas putida catechol 2,3-dioxygenase with high specific activity by genetically engineered Escherichia coli. J Biochem. 1995 Mar;117(3):614–622. doi: 10.1093/oxfordjournals.jbchem.a124753. [DOI] [PubMed] [Google Scholar]
  12. Koontz W. A., Shiman R. Beef kidney 3-hydroxyanthranilic acid oxygenase. Purification, characterization, and analysis of the assay. J Biol Chem. 1976 Jan 25;251(2):368–377. [PubMed] [Google Scholar]
  13. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  14. Nakai C., Hori K., Kagamiyama H., Nakazawa T., Nozaki M. Purification, subunit structure, and partial amino acid sequence of metapyrocatechase. J Biol Chem. 1983 Mar 10;258(5):2916–2922. [PubMed] [Google Scholar]
  15. Nishino S. F., Spain J. C. Degradation of nitrobenzene by a Pseudomonas pseudoalcaligenes. Appl Environ Microbiol. 1993 Aug;59(8):2520–2525. doi: 10.1128/aem.59.8.2520-2525.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Nozaki M., Kotani S., Ono K., Seno S. Metapyrocatechase. 3. Substrate specificity and mode of ring fission. Biochim Biophys Acta. 1970 Nov 11;220(2):213–223. doi: 10.1016/0005-2744(70)90007-0. [DOI] [PubMed] [Google Scholar]
  17. Okuno E., Köhler C., Schwarcz R. Rat 3-hydroxyanthranilic acid oxygenase: purification from the liver and immunocytochemical localization in the brain. J Neurochem. 1987 Sep;49(3):771–780. doi: 10.1111/j.1471-4159.1987.tb00960.x. [DOI] [PubMed] [Google Scholar]
  18. Parris G. E. Environmental and metabolic transformations of primary aromatic amines and related compounds. Residue Rev. 1980;76:1–30. doi: 10.1007/978-1-4612-6107-0_1. [DOI] [PubMed] [Google Scholar]
  19. Percival M. D. Human 5-lipoxygenase contains an essential iron. J Biol Chem. 1991 Jun 5;266(16):10058–10061. [PubMed] [Google Scholar]
  20. Que L., Jr Extradiol cleavage of o-aminophenol by pyrocatechase. Biochem Biophys Res Commun. 1978 Sep 14;84(1):123–129. doi: 10.1016/0006-291x(78)90272-3. [DOI] [PubMed] [Google Scholar]
  21. Shu L., Chiou Y. M., Orville A. M., Miller M. A., Lipscomb J. D., Que L., Jr X-ray absorption spectroscopic studies of the Fe(II) active site of catechol 2,3-dioxygenase. Implications for the extradiol cleavage mechanism. Biochemistry. 1995 May 23;34(20):6649–6659. doi: 10.1021/bi00020a010. [DOI] [PubMed] [Google Scholar]
  22. Stanier R. Y., Palleroni N. J., Doudoroff M. The aerobic pseudomonads: a taxonomic study. J Gen Microbiol. 1966 May;43(2):159–271. doi: 10.1099/00221287-43-2-159. [DOI] [PubMed] [Google Scholar]
  23. Stolz A., Nörtemann B., Knackmuss H. J. Bacterial metabolism of 5-aminosalicylic acid. Initial ring cleavage. Biochem J. 1992 Mar 15;282(Pt 3):675–680. doi: 10.1042/bj2820675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. VESCIA A., DI PRISCO G. Studies on purified 3-hydroxyanthranilic acid oxidase. J Biol Chem. 1962 Jul;237:2318–2324. [PubMed] [Google Scholar]
  25. Wolgel S. A., Dege J. E., Perkins-Olson P. E., Jaurez-Garcia C. H., Crawford R. L., Münck E., Lipscomb J. D. Purification and characterization of protocatechuate 2,3-dioxygenase from Bacillus macerans: a new extradiol catecholic dioxygenase. J Bacteriol. 1993 Jul;175(14):4414–4426. doi: 10.1128/jb.175.14.4414-4426.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES