The interactions of thiol compounds with porcine erythrocyte catalase.

Abstract

The effects of thiol compounds on the conformation of porcine erythrocyte catalase were examined. The thiol compounds showed two types of reactivity with the catalase in terms of changes in absorption spectra. One is characterized by the appearance of a new absorption maximum at 595 nm; this was seen with 2-mercaptoethanol (designated as inactive catalase type I). The other is characterized by new maxima at 535 and 570 nm, and this was seen with reduced glutathione, dithiothreitol, cysteine, and cysteamine (inactive catalase type II). The thiol compounds caused gradual inactivation of catalase, correlating with the enhancement of the absorption maximum at 595 nm or 570 nm. Removal of excess thiol reagents from the reaction mixtures caused partial recovery of activity, which was more marked with inactive catalase type II. Similar reversibility was observed in the absorption, CD and MCD spectra, whereas reversibility was not observed for inactive catalase type I. The MCD spectra suggested conversion of heme groups from a high to a low spin state on incubation with thiols, e.g., reduced glutathione, leading to inactive catalase type II. alpha-Helical conformation of the polypeptide backbone and titratable free SH groups in the catalase molecule were unaffected by all these thiol treatments. It is suggested that "active oxygen" which may be produced on incubation of catalase with thiol compounds, was responsible for the formation of inactive catalase type II.