The participation of a sulfhydryl group in the binding of iron to pyrocatechase

Abstract

Iron-free apopyrocatechase is easily converted to the holoenzyme containing one atom of ferric iron by treatment with ferrous iron under aerobic conditions. The presence of oxygen is essential for the reconstitution reaction and the stoichiometry among apoenzyme, ferrous iron and oxygen is 4 : 4 : 1. A sulfhydryl group of the apoenzyme, which reacts with either PCMB or DTNB, become undetectable in holoenzyme by DTNB. The apoenzyme modified with DTNB, loses its ability to bind iron unless treated with cysteine or β-mercaptoethanol. These results indicate that one free sulfhydryl group of the apoenzyme is involved in the binding of iron to the enzyme through iron-sulfur bond and show the possibility of the participation of an oxidase-like reaction in the reconstitution of holoenzyme.