Metapyrocatechase

Abstract

Three times crystallized metapyrocatechase has a specific activity of about 110 pmoles per mln per mg of protein at 24" and contains 1 g atom of iron per mole of enzyme, based on molecular weight of 140,000.The amino acid composition of the enzyme is presented.Metapyrocatechase is easily inactivated by various oxidizing agents, such as air or HzOz.This inactivation appears to be due to oxidation of ferrous ion to the ferric form, which is then released from the enzyme protein.The enzyme thus inactivated can be fully reactivated by incubation with ferrous ion and a reducing agent under anaerobic conditions.Inactivation of the enzyme is prevented by the presence of substrate, catechol.Enzyme activity is inhibited by a variety of nitrogencontaining aromatic compounds, including o-phenanthroline, CY, or'dipyridyl, m-phenanthroline, a-naphthoquinollne, quinoline, and pyridine.Inhibition by these agents is shown to be competitive with respect to the substrate, catechol.On the other hand, these compounds protect the enzyme from inactivation by oxidizing agents, and prevent reactivation of inactivated enzyme.Organic solvents, such as acetone or ethanol, also protect the enzyme from inactivation, and inhibit the enzyme activity nearly competitively with respect to catechol.Metapyrocatechase contains 4 free sulfhydryl groups per mole of enzyme out of 12 total half-cystine residues.When the 4 free sulfhydryl groups are titrated by p-chloromercuribenzoate, no loss of enzymatic activity is observed.However, the enzyme is inactivated by an excess amount of