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Open Access Highly Accessed Research article

Mechanisms underlying dioxygen reduction in laccases. Structural and modelling studies focusing on proton transfer

Isabel Bento*, Catarina S Silva, Zhenjia Chen, Lígia O Martins, Peter F Lindley and Cláudio M Soares*

Author Affiliations

Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Av. da República, 2780-157 Oeiras, Portugal

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BMC Structural Biology 2010, 10:28  doi:10.1186/1472-6807-10-28

Published: 7 September 2010

Abstract

Background

Laccases are enzymes that couple the oxidation of substrates with the reduction of dioxygen to water. They are the simplest members of the multi-copper oxidases and contain at least two types of copper centres; a mononuclear T1 and a trinuclear that includes two T3 and one T2 copper ions. Substrate oxidation takes place at the mononuclear centre whereas reduction of oxygen to water occurs at the trinuclear centre.

Results

In this study, the CotA laccase from Bacillus subtilis was used as a model to understand the mechanisms taking place at the molecular level, with a focus in the trinuclear centre. The structures of the holo-protein and of the oxidised form of the apo-protein, which has previously been reconstituted in vitro with Cu(I), have been determined. The former has a dioxygen moiety between the T3 coppers, while the latter has a monoatomic oxygen, here interpreted as a hydroxyl ion. The UV/visible spectra of these two forms have been analysed in the crystals and compared with the data obtained in solution. Theoretical calculations on these and other structures of CotA were used to identify groups that may be responsible for channelling the protons that are needed for reduction of dioxygen to water.

Conclusions

These results present evidence that Glu 498 is the only proton-active group in the vicinity of the trinuclear centre. This strongly suggests that this residue may be responsible for channelling the protons needed for the reduction. These results are compared with other data available for these enzymes, highlighting similarities and differences within laccases and multicopper oxidases.