Reductant-dependent electron distribution among redox sites of laccase.

Abstract

Rhus vernicifera laccase (monophenol monooxygenase, monophenol, dihydroxyphenylalanine:oxygen oxidoreductase, EC 1.14.18.1) an .**GRAPHIC**. oxidoreductase containing 4 Cu ions bound to 3 redox sites (type 1, type 2 and type 3 Cu pair), was titrated anaerobically with several reductants having various chemical and thermodynamic properties. The distribution of electron equivalents among the redox sites was found to be reductant dependent. When the data for titration by various reductants of the type 3 site were plotted against those of the type 1 site according to the Nernst formalism, the slope n varied from 2.0-1.0. The redox potential of the reductant''s 1st oxidation step is qualitatively correlated with the value of n and is suggested as the factor that modulates the electron distribution. Such a behavior implies a nonequilibrium situation. A very good simulation of the data was provided by an analysis assuming a formally variable cooperativity between the 2 type 3 Cu ions. This apparent variability is suggested to result from a process whereby sufficiently strong reductants induce a transition of the type 3 site from a cooperative 2-electron acceptor to a pair of independent 1-electron acceptors. This uncoupled state of the type 3 site is considered metastable. Other possible models were also investigated. Apparently the 2-electron accepting behavior of the 330 nm chromophore is the exception rather than the rule.