Metapyrogatechase: Catechol Oxidation in Azotobacter Vinelandii

Benzoic acid metabolism was studied in Azotobacter vinelandii. 14 A radioactive intermediate of C-benzoic acid metabolism was discovered. Preliminary evidence suggests that this intermediate may be 2 ,-hydroxybenzoate but positive identification was not achieved, values for several hydroxybenzoates and catechol on paper chromato­ graphy in various solvent systems are given. The oxygenase, metapyrocatechase, which catalyzes the oxida­ tive cleavage of catechol to form a-hydroxy-cis, cis-muconic semi­ aldehyde, was purified about 2 0 -fold from the soluble cell extract of benzoate grown cells, and partially characterized. The enzyme contains two sulfhydryl groups, readily titrable with 5,5 *-dithio-bis(2 -nitrobenzoic acid), which are not necessary for enzyme activity, + -4 4-5Ag (1 x 10 M) completely inactivated the enzyme while Hg (1 x -4 10 M) only partially inactivated it, Thioglycollate reactivated -j-| . inactive enzyme, while Fe had no reactivating effect. -6 The for catechol is 1.7 x 10 M. Azotobacter metapyrocatechase uses the following compounds as substrates t catechol, 3-methyl catechol, 4-methyl catechol, 3,4-dihydroxyphenylacetate, 3,4-dihydroxybenzoate, DL-3,4-dihydroxyphenylalanine, 3,4-dihydroxyhydrocinnamate, and 3,4-dihydroxycinnamate. It does not use 2,3-dihydroxybenzoate, £-aminophenol, or guaiacol. . High concentrations of catechol, 3-methyl catechol, or 4-methyl catechol inhibit enzyme activity. Thus