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J. Biol. Chem., Vol. 281, Issue 52, 40264-40272, December 29, 2006

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Atomic Level Insight into the Oxidative Half-reaction of Aromatic Amine Dehydrogenase^*

From the Manchester Interdisciplinary Biocentre, University of Manchester, Manchester M1 7DN, United Kingdom

The quinoprotein aromatic amine dehydrogenase (AADH) uses a covalently bound tryptophan tryptophylquinone (TTQ) cofactor to oxidatively deaminate primary aromatic amines. Recent crystal structures have provided insight into the reductive half-reaction. In contrast, no atomic details are available for the oxidative half-reaction. The TTQ O7 hydroxyl group is protonated during reduction, but it is unclear how this proton can be removed during the oxidative half-reaction. Furthermore, compared with the electron transfer from the N-quinol form, electron transfer from the non-physiological O-quinol form to azurin is significantly slower. Here we report crystal structures of the O-quinol, N-quinol, and N-semiquinone forms of AADH. A comparison of oxidized and substrate reduced AADH species reveals changes in the TTQ-containing subunit, extending from residues in the immediate vicinity of the N-quinol to the putative azurin docking site, suggesting a mechanism whereby TTQ redox state influences interprotein electron transfer. In contrast, chemical reduction of the TTQ center has no significant effect on protein conformation. Furthermore, structural reorganization upon substrate reduction places a water molecule near TTQ O7 where it can act as proton acceptor. The structure of the N-semiquinone, however, is essentially similar to oxidized AADH. Surprisingly, in the presence of substrate a covalent N-semiquinone substrate adduct is observed. To our knowledge this is the first detailed insight into a complex, branching mechanism of quinone oxidation where significant structural reorganization upon reduction of the quinone center directly influences formation of the electron transfer complex and nature of the electron transfer process.

Received for publication, June 9, 2006 , and in revised form, September 11, 2006.

The atomic coordinates and structure factors (code 2iur, 2iuv, 2iup, 2iuq, and 2hxc) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).

^* The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at http://www.jbc.org) contains supplemental Fig. 1.

¹ To whom correspondence should be addressed: Manchester Interdisciplinary Biocentre, University of Manchester, Princess St. 131, Manchester, M1 7ND, UK. Tel.: 44-161-306-51-50; E-mail: david.leys{at}manchester.ac.uk.

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This article has been cited by other articles:

				A. Roujeinikova, P. Hothi, L. Masgrau, M. J. Sutcliffe, N. S. Scrutton, and D. Leys New Insights into the Reductive Half-reaction Mechanism of Aromatic Amine Dehydrogenase Revealed by Reaction with Carbinolamine Substrates J. Biol. Chem., August 17, 2007; 282(33): 23766 - 23777. [Abstract] [Full Text] [PDF]