On the Possibility of Uphill Intramolecular Electron Transfer in Multicopper Oxidases: Electrochemical and Quantum Chemical Study of Bilirubin Oxidase |
| |
Authors: | Sergey Shleev Viktor Andoralov Magnus Falk Curt T Reimann Tautgirdas Ruzgas Martin Srnec Ulf Ryde Lubomír Rulíšek |
| |
Institution: | 1. The Laboratory of Chemical Enzymology, A.?N. Bach Institute of Biochemistry RAS, 119071 Moscow, Russia fax: +7?495?954?2732;2. The Department of Biomedical Science, Malm? University, 205?06 Malm?, Sweden;3. The laboratory of Electrocatalysis and Fuel Cells, A.?N. Frumkin Institute of Physical Chemistry and Electrochemistry RAS, 119071 Moscow, Russia;4. Department of Pure and Applied Biochemistry, Lund University, 22100 Lund, Sweden;5. Institute of Organic Chemistry and Biochemistry, Gilead Sciences Research Center at IOCB, Academy of Sciences of the Czech Republic, 166?10 Praha 6, Czech Republic;6. Department of Theoretical Chemistry, Lund University, 22100 Lund, Sweden |
| |
Abstract: | The catalytic cycle of multicopper oxidases (MCOs) involves intramolecular electron transfer (IET) from the Cu‐T1 copper ion, which is the primary site of the one‐electron oxidations of the substrate, to the trinuclear copper cluster (TNC), which is the site of the four‐electron reduction of dioxygen to water. In this study we report a detailed characterization of the kinetic and electrochemical properties of bilirubin oxidase (BOx) – a member of the MCO family. The experimental results strongly indicate that under certain conditions, e.g. in alkaline solutions, the IET can be the rate‐limiting step in the BOx catalytic cycle. The data also suggest that one of the catalytically relevant intermediates (most likely characterized by an intermediate oxidation state of the TNC) formed during the catalytic cycle of BOx has a redox potential close to 0.4 V, indicating an uphill IET process from the T1 copper site (0.7 V) to the Cu‐T23. These suggestions are supported by calculations of the IET rate, based on the experimentally observed Gibbs free energy change and theoretical estimates of reorganization energy obtained by combined quantum and molecular mechanical (QM/MM) calculations. |
| |
Keywords: | Bilirubin oxidase Intramolecular electron transfer Rate‐limiting catalytic step Reorganization energy QM/MM calculations |
|
|