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J Am Chem Soc. 2020 Jan 22;142(3):1394-1405. doi: 10.1021/jacs.9b11147. Epub 2020 Jan 8.

Interplay between Orientation at Electrodes and Copper Activation of Thermus thermophilus Laccase for O2 Reduction.

Author information

1
Aix Marseille Univ, CNRS, BIP UMR 7281 , 31 Chemin Aiguier , CS 70071, 13402 Marseille , Cedex 09 , France.
2
Aix Marseille Univ, CNRS, IMM FR 3479 , 31 Chemin Aiguier , CS 70071, 13402 Marseille , Cedex 09 , France.
3
Aix Marseille Univ, CNRS, CINAM UMR 7325, Campus de Luminy , 13288 Marseille , Cedex 09 , France.
4
Aix Marseille Univ, Université de Toulon, CNRS, IM2NP UMR 7334 , 13397 Marseille , France.

Abstract

Multicopper oxidases (MCOs) catalyze the oxidation of a variety of substrates while reducing oxygen into water through four copper atoms. As an additional feature, some MCOs display an enhanced activity in solution in the presence of Cu2+. This is the case of the hyperthermophilic laccase HB27 from Thermus thermophilus, the physiologic role of which is unknown. As a particular feature, this enzyme presents a methionine rich domain proposed to be involved in copper interaction. In this work, laccase from T. thermophilus was produced in E. coli, and the effect of Cu2+ on its electroactivity at carbon nanotube modified electrodes was investigated. Direct O2 electroreduction is strongly dictated by carbon nanotube surface chemistry in accordance with the enzyme dipole moment. In the presence of Cu2+, an additional low potential cathodic wave occurs, which was never described earlier. Analysis of this wave as a function of Cu2+ availability allows us to attribute this wave to a cuprous oxidase activity displayed by the laccase and induced by copper binding close to the Cu T1 center. A mutant lacking the methionine-rich hairpin domain characteristic of this laccase conserves its copper activity suggesting a different site of copper binding. This study provides new insight into the copper effect in methionine rich MCOs and highlights the utility of the electrochemical method to investigate cuprous oxidase activity and to understand the physiological role of these MCOs.

PMID:
31865707
DOI:
10.1021/jacs.9b11147

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