Spectroscopic Investigation of Peroxide Binding to the Trinuclear Copper Cluster Site in Laccase: Correlation with the Peroxy-Level Intermediate and Relevance to Catalysis

Abstract

Laccase is a multicopper oxidase which contains four coppers, one type 1, one type 2, and a coupled binuclear type 3 pair, the type 2 and type 3 copper centers together forming a trinuclear copper cluster. The type 1 mercury derivative of laccase (T1Hg Lc) has the type 1 center substituted with a redox-inactive Hg2+ ion and an intact trinuclear copper cluster. Reaction of H2O2 with fully oxidized T1Hg Lc produces a peroxide adduct which has now been studied in detail. Peroxide is shown to bind to the trinuclear cluster with low affinity, producing spectral and geometric features similar to the intermediate formed in the reduction of O2 to H2O which had been shown to have the type 2 copper reduced, the type 3 pair oxidized and antiferromagnetically coupled, and two coppers bridged at a distance of 3.4 A. The peroxide adduct and the intermediate have similar geometric and electronic features with the type 2 oxidized in the adduct. This center is paramagnetic and has been studied in detail. Peroxide binds to...