EVIDENCE FOR AN UNUSUAL ELECTRONIC-STRUCTURE OF WHEAT-GERM PEROXIDASECOMPOUND-I

Oxidized states of wheat germ peroxidase isozyme C2 (WGP C2) were inve stigated by means of electronic absorption spectroscopy. Addition of o ne molar equivalent of H2O2 to ferric WGP C2 led to the formation of a n oxidized species with an absorption spectrum very similar to that of peroxidase compound II, with a Soret maximum at 411 nm and visible ma xima at 523 and 553 nm, The transformation took place with an isosbest ic point at 409 nm, Stopped flow spectroscopy showed no inflection poi nts for the formation of this species when it was registered at 420 nm , and we could verify the persistence of the isosbestic point from 20 ms to 10 s, The oxidized species decays spontaneously to ferric enzyme in a double-exponential manner. By adding excess H2O2 to the system w e obtained an inactive derivative identical to horseradish peroxidase P-670, In the presence of one equivalent of reducing substrate and exc ess H2O2 compound III was formed. The results so indicate that the spe cies obtained in the reaction of WGP C2 with equimolecular amounts of H2O2 is compound I. The resulting compound I spectrum was identical to that of cytochrome c peroxidase, suggesting the formation of a protei n radical rather than the typical pi cation radical, a feature which h ad not been described before for a plant peroxidase. (C) 1998 Academic Press.