2-AMINOPHENOL 1,6-DIOXYGENASE - A NOVEL AROMATIC RING-CLEAVAGE ENZYMEPURIFIED FROM PSEUDOMONAS PSEUDOALCALIGENES JS45

Most bacterial pathways for the degradation of aromatic compounds invo lve introduction of two hydroxyl groups either ortho or para to each o ther, Ring fission then occurs at the bond adjacent to one of the hydr oxyl groups, In contrast, 2-aminophenol is cleaved to 2-aminomuconic a cid semialdehyde in the nitrobenzene-degrading strain Pseudomonas pseu doalcaligenes JS45. To examine the relationship between this enzyme an d other dioxygenases, 2-aminophenol 1,6-dioxygenase has been purified by ethanol precipitation, gel filtration, and ion exchange chromatogra phy. The molecular mass determined by gel filtration was 140,000 Da, S odium dodecyl sulfate-polyacrylamide gel electrophoresis showed two su bunits of 35,000 and 39,000 Da, which suggested an alpha(2) beta(2) su bunit structure, Studies with inhibitors indicated that ferrous iron w as the sole cofactor, The K-m values for 2-aminophenol and oxygen were 4.2 and 710 mu M, respectively, The enzyme catalyzed the oxidation of catechol, 6-amino-m-cresol, 2-amino-m-cresol, and 2-amino-4-chlorophe nol. 3-Hydroxyanthranilate, protocatechuate, gentisate, and 3- and 4-m ethylcatechol were not substrates. The substrate range and the subunit structure are unique among those of the known ring cleavage dioxygena ses.