Functional identification of the gene locus ncg12319 and characterization of catechol 1,2-dioxygenase in Corynebacterium glutamicum

Corynebacterium glutamicum assimilated phenol, benzoate, 4-hydroxybenzoate p-cresol and 3,4-dihydroxybenzoate. Ring cleavage was by catechol 1,2-dioxygenase when phenol or benzoate was used and by protocatechuate 3,4-dioxygenase when the others were used as substrate. The locus ncg12319 of its genome was cloned and expressed in Escherichia coli. Enzyme assays showed that ncg12319 encodes a catechol 1,2-dioxygenase. This catechol 1,2-dioxygenase was purified and accepted catechol, 3-, or 4-methylcatechols, but not chlorinated catechols, as substrates. The optimal temperature and pH for catechol cleavage catalyzed by the enzyme were 30°C and 9, respectively, and the Km and Vmax were determined to be 4.24 μmol 1 -1 and 3.7 μmol 1-1 min-1 mg-1 protein, respectively.