Pseudomonas aeruginosa 142 uses a three-component ortho-halobenzoate 1,2-dioxygenase for metabolism of 2,4-dichloro- and 2-chlorobenzoate

J Bacteriol. 1994 Jun;176(11):3368-74. doi: 10.1128/jb.176.11.3368-3374.1994.

Abstract

Cell extracts of Pseudomonas aeruginosa 142, which was previously isolated from a polychlorinated biphenyl-degrading consortium, were shown to degrade 2,4-dichlorobenzoate, 2-chlorobenzoate, and a variety of other substituted ortho-halobenzoates by a reaction that requires oxygen, NADH, Fe(II), and flavin adenine dinucleotide. By using extracts that were chromatographically depleted of chlorocatechol and catechol 1,2-dioxygenase activities, products of the initial reaction with 2,4- or 2,5-dichlorobenzoate and 2-chlorobenzoate were identified by mass spectrometry as 4-chlorocatechol and catechol. In contrast to the well-characterized benzoate dioxygenases or the recently described 2-halobenzoate 1,2-dioxygenase from P. cepacia 2CBS (S. Fetzner, R. Müller, and F. Lingens, J. Bacteriol. 174:279-290, 1992) that possess two protein components, the P. aeruginosa enzyme was resolved by ion-exchange chromatography into three components, each of which is required for activity. To verify the distinct nature of this enzyme, we purified, characterized, and identified one component as a ferredoxin (M(r), approximately 13,000) containing a single [2Fe-2S] Rieske-type cluster (electron paramagnetic resonance spectroscopic values of gx = 1.82, gy = 1.905, and gz = 2.02 in the reduced state) that is related in sequence to ferredoxins found in the naphthalene and biphenyl three-component dioxygenase systems. By analogy to these enzymes, we propose that the P. aeruginosa ferredoxin serves as an electron carrier between an NADH-dependent ferredoxin reductase and the terminal component of the ortho-halobenzoate 1,2-dioxygenase. The broad specificity and high regiospecificity of the enzyme make it a promising candidate for use in the degradation of mixtures of chlorobenzoates.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Catechols / metabolism
  • Chlorobenzoates / metabolism*
  • Molecular Sequence Data
  • Oxidation-Reduction
  • Oxygenases / isolation & purification
  • Oxygenases / metabolism*
  • Pseudomonas aeruginosa / enzymology
  • Pseudomonas aeruginosa / metabolism*
  • Sequence Analysis
  • Sequence Homology, Amino Acid
  • Substrate Specificity

Substances

  • Catechols
  • Chlorobenzoates
  • 2,4-dichlorobenzoic acid
  • 2-chlorobenzoic acid
  • Oxygenases
  • ortho-halobenzoate 1,2-dioxygenase
  • 4-chlorocatechol
  • catechol