Purification and some properties of component A of the 4-chlorophenylacetate 3,4-dioxygenase from Pseudomonas species strain CBS

J Biol Chem. 1986 Sep 25;261(27):12883-8.


Pseudomonas sp. strain CBS 3 possesses a two-component enzyme system which converts 4-chlorophenylacetate to 3,4-dihydroxyphenylacetate by the incorporation of 2 atoms of molecular oxygen. Component A of this enzyme system was purified to homogeneity by a 5-step procedure. After the last purification step the enzyme was homogeneous in analytical and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The molecular weight of the native protein was determined to be 140,000 by Sephadex G-200 and 144,000 by analytical ultracentrifugation. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that component A consists of three identical subunits with a molecular weight determined to range between 46,000 and 52,000. The isoelectric point was estimated to be 5.0. Component A shows an intensive red-brown color, and in the oxidized state it exhibits a visible absorption spectrum with a maximum at 458 nm and a shoulder at 560 nm. By reduction with sodium dithionite a new peak with a maximum at 518-520 nm is observed. The enzyme contains iron (1.6-1.8 mol/subunit) and acid-labile sulfide (1.6-1.9 mol/subunit) which suggests that component A is an iron-sulfur protein.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids / analysis
  • Chromatography, Affinity
  • Chromatography, Gel
  • Chromatography, Ion Exchange
  • Dioxygenases*
  • Isoelectric Point
  • Molecular Weight
  • Oxygenases / isolation & purification*
  • Pseudomonas / enzymology*


  • Amino Acids
  • Oxygenases
  • 4-chlorophenylacetate 3,4-dioxygenase
  • Dioxygenases