A New Electron Transport Mechanism in Mitochondrial Steroid Hydroxylase Systems Based on Structural Changes upon the Reduction of Adrenodoxin

The adrenal ferredoxin (adrenodoxin, Adx) is an acidic 14.4-kDa [2Fe-2S] ferredoxin that belongs to the vertebrate ferredoxin family. It is involved in the electron transfer from the flavoenzyme NADPH-adrenodoxin-reductase to cytochromes P-450scc and P-45011 β. The interaction between the redox part...

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Bibliographic Details
Published in:Biochemistry (Easton) 2002-06, Vol.41 (25), p.7969-7978
Main Authors: Beilke, Dirk, Weiss, Roland, Löhr, Frank, Pristovšek, Primož, Hannemann, Frank, Bernhardt, Rita, Rüterjans, Heinz
Format: Article
Language:English
Subjects:
Adrenodoxin - chemistry
Adrenodoxin - metabolism
Animals
Cattle
Computer Simulation
Cytochrome P-450 Enzyme System - chemistry
Cytochrome P-450 Enzyme System - metabolism
Dimerization
Electron Transport
Ferredoxin-NADP Reductase - chemistry
Ferredoxin-NADP Reductase - metabolism
Mitochondria - enzymology
Models, Molecular
Nuclear Magnetic Resonance, Biomolecular
Oxidation-Reduction
Protein Structure, Secondary
Protein Structure, Tertiary
Steroid Hydroxylases - chemistry
Steroid Hydroxylases - metabolism
Structure-Activity Relationship
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Summary:The adrenal ferredoxin (adrenodoxin, Adx) is an acidic 14.4-kDa [2Fe-2S] ferredoxin that belongs to the vertebrate ferredoxin family. It is involved in the electron transfer from the flavoenzyme NADPH-adrenodoxin-reductase to cytochromes P-450scc and P-45011 β. The interaction between the redox partners during electron transport has not yet been fully established. Determining the tertiary structure of an electron-transfer protein may be very helpful in understanding the transport mechanism. In the present work, we report a structural study on the oxidized and reduced forms of bovine adrenodoxin (bAdx) in solution using high-resolution NMR spectroscopy. The protein was produced in Escherichia coli and singly or doubly labeled with 15N or 13C/15N, respectively. Approximately 70 and 75% of the 15N, 13C, and 1H resonances could be assigned for the reduced and the oxidized bAdx, respectively. The secondary and tertiary structures of the reduced and oxidized states were determined using NOE distance information. 1HN-T1 relaxation times of certain residues were used to obtain additional distance constraints to the [2Fe-2S] cluster. The results suggest that the solution structure of oxidized Adx is quite similar to the X-ray structure. However, structural changes occur upon reduction of the [2Fe-2S] cluster, as indicated by NMR measurements. It could be shown that these conformational changes, especially in the C-terminal region, cause the dissociation of the Adx dimer upon reduction. A new electron transport mechanism proceeding via a modified shuttle mechanism, with both monomers and dimers acting as electron carriers, is proposed.
ISSN:0006-2960
1520-4995
DOI:10.1021/bi0160361