Concerted electron/proton transfer mechanism in the oxidation of phenols by laccase

This study aimed to assess structural requirements in the enzyme/substrate interactions that are responsible for tuning the enzymatic reactivity. To better assess the role of the aspartic residue in the substrate-binding pocket of basidiomycete-type laccases, we compared the catalytic efficiency of wild-type enzymes to that of a mutant in which carboxylic acid residue Asp206 was changed to alanine. Oxidation efficiency towards phenolic substrates by laccases of Trametes villosa, Trametes versicolor and a T. versicolor D206A mutant was studied at two pH values. By the Hammett approach and Marcus analysis, we obtained unambiguous evidence that the oxidation takes place by a concerted electron/proton transfer (EPT) mechanism, and that at pH 5 (optimum pH for enzyme activity) the phenolic proton is transferred to Asp206 during the concerted electron/proton transfer process. Oxidation efficiency towards phenolic substrates by Trametes villosa and Trametes versicolor laccases and the D206A mutant of this latter was studied at two pH values. By the Hammett approach and Marcus analysis, we obtained unambiguous evidence that the oxidation takes place by a concerted electron/proton transfer (EPT) mechanism. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.