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Article

Light-Induced Reduction of Bovine Adrenodoxin via the Covalently Bound Ruthenium(II) Bipyridyl Complex: Intramolecular Electron Transfer and Crystal Structure^†

Supporting Info

Andrei Halavaty,^‡ Jürgen J. Müller,^‡ Jörg Contzen,^§ Christiane Jung,^§ Frank Hannemann, Rita Bernhardt, Marcus Galander, Friedhelm Lendzian, and Udo Heinemann*^‡^#

Crystallography Group and Protein Dynamics, Max-Delbrck-Centrum fr Molekulare Medizin, Robert-Rssle-Strasse 10, D-13125 Berlin-Buch, Germany, Department of Biochemistry, Universitt des Saarlandes, Postfach 15 11 50, D-66041 Saarbrcken, Germany, Max-Volmer-Laboratorium fr Biophysikalische Chemie, Institut fr Chemie, Technische Universitt Berlin, PC 14, Strasse des 17. Juni 135, D-10623 Berlin, Germany, and Institut fr Chemie/Kristallographie, Freie Universitt, Takustrasse 6, D-14195 Berlin, Germany

Biochemistry, 2006, 45 (3), pp 709–718

DOI: 10.1021/bi0510330

Publication Date (Web): December 22, 2005

†

This work was supported by the DFG, Projects He 1318/19-3.4 and Ju 229/4-3.

‡

Crystallography Group, Max-Delbrück-Centrum für Molekulare Medizin.

Protein Dynamics, Max-Delbrück-Centrum für Molekulare Medizin.

Universität des Saarlandes.

Technische Universität Berlin.

To whom correspondence should be addressed at the Crystallography Group, Max-Delbrück-Centrum für Molekulare Medizin. Phone: (49 30) 94063420. Fax: (49 30) 94062548. E-mail: heinemann@mdc-berlin.de.

Freie Universität.

Abstract

Bovine adrenodoxin (Adx) plays an important role in the electron-transfer process in the mitochondrial steroid hydroxylase system of the bovine adrenal cortex. Using electron paramagnetic resonance (EPR) spectroscopy, we showed that photoreduction of the [2Fe-2S] cluster of Adx via (4‘-methyl-2,2‘-bipyridine)bis(2,2‘-bipyridine)ruthenium(II) [Ru(bpy)₂(mbpy)] covalently attached to the protein surface can be used as a new approach to probe the “shuttle” hypothesis for the electron transfer by Adx. The 1.5 Å resolution crystal structure of a 1:1 Ru(bpy)₂(mbpy)−Adx(1−108) complex reveals the site of modification, Cys95, and allows to predict the possible intramolecular electron-transfer pathways within the complex. Photoreduction of uncoupled Adx, mutant Adx(1−108), and Ru(bpy)₂(mbpy)−Adx(1−108) using safranin T as the mediating electron donor suggests that two electrons are transferred from the dye to Adx. The intramolecular photoreduction rate constant for the ruthenated Adx has been determined and is discussed according to the predicted pathways.

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