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. 1978 Apr;134(1):30–37. doi: 10.1128/jb.134.1.30-37.1978

Purification and properties of 2-hydroxy-6-oxo-2,4-heptadienoate hydrolase from two strains of Pseudomonas putida.

R C Bayly, D di Berardino
PMCID: PMC222214  PMID: 77272

Abstract

Growth on phenol of two strains of Pseudomonas putida biotype A, NCIB 10015 and NCIB 9865, elicits the synthesis of an enzyme that hydrolyzes 2-hydroxy-6-oxo-2,4-heptadienoate to 2-oxopent-4-enoate. The purified enzyme from Pseudomonas NCIB 10015 has a molecular weight of 118,000 and dissociates in sodium dodecyl sulfate to a species of molecular weight 27,700; the enzyme from Pseudomonas NCIB 9865 has a molecular weight of 100,000 and dissociates to a species of 25,000 molecular weight. The hydrolases from both strains have similar Km values, pH optima, and thermal labilities and attack the same range of substrates. Neither hydrolase was stimulated by Mg2+ or Mn2+, and both were inhibited by p-chloromercuribenzoate and iodoacetamide. Immunodiffusion studies with the purified enzymes and antibodies formed against them show some cross-reaction of Pseudomonas NCIB 9865 enzymes with antibodies to Pseudomonas NCIB 10015, but not vice versa.

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Selected References

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  1. Andrews P. Estimation of the molecular weights of proteins by Sephadex gel-filtration. Biochem J. 1964 May;91(2):222–233. doi: 10.1042/bj0910222. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bayly R. C., Dagley S., Gibson D. T. The metabolism of cresols by species of Pseudomonas. Biochem J. 1966 Nov;101(2):293–301. doi: 10.1042/bj1010293. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bayly R. C., Dagley S. Oxoenoic acids as metabolites in the bacterial degradation of catechols. Biochem J. 1969 Feb;111(3):303–307. doi: 10.1042/bj1110303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bayly R. C., Wigmore G. J. Metabolism of phenol and cresols by mutants of Pseudomonas putida. J Bacteriol. 1973 Mar;113(3):1112–1120. doi: 10.1128/jb.113.3.1112-1120.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Clarke P. H. Biochemical and immunological comparison of aliphatic amidases produced by Pseudomonas species. J Gen Microbiol. 1972 Jul;71(2):241–257. doi: 10.1099/00221287-71-2-241. [DOI] [PubMed] [Google Scholar]
  6. Collinsworth W. L., Chapman P. J., Dagley S. Stereospecific enzymes in the degradation of aromatic compounds by pseudomonas putida. J Bacteriol. 1973 Feb;113(2):922–931. doi: 10.1128/jb.113.2.922-931.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Crawford R. L., Hutton S. W., Chapman P. J. Purification and properties of gentisate 1,2-dioxygenase from Moraxella osloensis. J Bacteriol. 1975 Mar;121(3):794–799. doi: 10.1128/jb.121.3.794-799.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. DAVIS B. J. DISC ELECTROPHORESIS. II. METHOD AND APPLICATION TO HUMAN SERUM PROTEINS. Ann N Y Acad Sci. 1964 Dec 28;121:404–427. doi: 10.1111/j.1749-6632.1964.tb14213.x. [DOI] [PubMed] [Google Scholar]
  9. Dagley S. A biochemical approach to some problems of environmental pollution. Essays Biochem. 1975;11:81–138. [PubMed] [Google Scholar]
  10. Hayaishi O. Crystalline oxygenases of pseudomonads. Bacteriol Rev. 1966 Dec;30(4):720–731. doi: 10.1128/br.30.4.720-731.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Johnson B. F., Stanier R. Y. Dissimilation of aromatic compounds by Alcaligenes eutrophus. J Bacteriol. 1971 Aug;107(2):468–475. doi: 10.1128/jb.107.2.468-475.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  13. Leung P. T., Chapman P. J., Dagley S. Purification and properties of 4-hydroxy-2-ketopimelate aldolase from Acinetobacter. J Bacteriol. 1974 Oct;120(1):168–172. doi: 10.1128/jb.120.1.168-172.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Meagher R. B., Ornston L. N. Relationships among enzymes of the beta-ketoadipate pathway. I. Properties of cis,cis-muconate-lactonizing enzyme and muconolactone isomerase from Pseudomonas putida. Biochemistry. 1973 Aug 28;12(18):3523–3530. doi: 10.1021/bi00742a027. [DOI] [PubMed] [Google Scholar]
  15. Ribbons D. W. Metabolism of omicron-cresol by Pseudomonas aeruginosa strain T1. J Gen Microbiol. 1966 Aug;44(2):221–231. doi: 10.1099/00221287-44-2-221. [DOI] [PubMed] [Google Scholar]
  16. Sala-Trepat J. M., Evans W. C. The meta cleavage of catechol by Azotobacter species. 4-Oxalocrotonate pathway. Eur J Biochem. 1971 Jun 11;20(3):400–413. doi: 10.1111/j.1432-1033.1971.tb01406.x. [DOI] [PubMed] [Google Scholar]
  17. Sala-Trepat J. M., Murray K., Williams P. A. The metabolic divergence in the meta cleavage of catechols by Pseudomonas putida NCIB 10015. Physiological significance and evolutionary implications. Eur J Biochem. 1972 Jul 24;28(3):347–356. doi: 10.1111/j.1432-1033.1972.tb01920.x. [DOI] [PubMed] [Google Scholar]
  18. Sparnins V. L., Chapman P. J., Dagley S. Bacterial degradation of 4-hydroxyphenylacetic acid and homoprotocatechuic acid. J Bacteriol. 1974 Oct;120(1):159–167. doi: 10.1128/jb.120.1.159-167.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. 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]
  20. Stanier R. Y., Wachter D., Gasser C., Wilson A. C. Comparative immunological studies of two Pseudomonas enzymes. J Bacteriol. 1970 May;102(2):351–362. doi: 10.1128/jb.102.2.351-362.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Tack B. F., Chapman P. J., Dagley S. Purification and properties of 4-hydroxy-4-methyl-2-oxoglutarate aldolase. J Biol Chem. 1972 Oct 25;247(20):6444–6449. [PubMed] [Google Scholar]
  22. Tai H. H., Sih C. J. 3,4-dihydroxy-9,10-secoandrosta-1,3,5(10)-triene-9,17-dione 4,5-dioxygenase from Nocardia restrictus. I. Isolation of the enzyme and study of its physical and chemical properties. J Biol Chem. 1970 Oct 10;245(19):5062–5071. [PubMed] [Google Scholar]
  23. Weber K., Osborn M. The reliability of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J Biol Chem. 1969 Aug 25;244(16):4406–4412. [PubMed] [Google Scholar]
  24. Wigmore G. J., Bayly R. C., Di Berardino D. Pseudomonas putida mutants defective in the metabolism of the products of meta fission of catechol and its methyl analogues. J Bacteriol. 1974 Oct;120(1):31–37. doi: 10.1128/jb.120.1.31-37.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]

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