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Structural biology of redox partner interactions in P450cam monooxygenase: A fresh look at an old system

Authors
  • Sevrioukova, Irina F.
  • Poulos, Thomas L.
Type
Published Article
Journal
Archives of Biochemistry and Biophysics
Publisher
Elsevier BV
Publication Date
Jan 01, 2010
Volume
507
Issue
1
Pages
66–74
Identifiers
DOI: 10.1016/j.abb.2010.08.022
Source
Elsevier
Keywords
License
Unknown

Abstract

The P450cam monooxygenase system consists of three separate proteins: the FAD-containing, NADH-dependent oxidoreductase (putidaredoxin reductase or Pdr), cytochrome P450cam and the 2Fe2S ferredoxin (putidaredoxin or Pdx), which transfers electrons from Pdr to P450cam. Over the past few years our lab has focused on the interaction between these redox components. It has been known for some time that Pdx can serve as an effector in addition to its electron shuttle role. The binding of Pdx to P450cam is thought to induce structural changes in the P450cam active site that couple electron transfer to substrate hydroxylation. The nature of these structural changes has remained unclear until a particular mutant of P450cam (Leu358Pro) was found to exhibit spectral perturbations similar to those observed in wild type P450cam bound to Pdx. The crystal structure of the L358P variant has provided some important insights on what might be happening when Pdx docks. In addition to these studies, many Pdx mutants have been analyzed to identify regions important for electron transfer. Somewhat surprisingly, we found that Pdx residues predicted to be at the P450cam–Pdx interface play different roles in the reduction of ferric P450cam and the ferrous P450–O 2 complex. More recently we have succeeded in obtaining the structure of a chemically cross-linked Pdr–Pdx complex. This fusion protein represents a valid model for the noncovalent Pdr–Pdx complex as it retains the redox activities of native Pdr and Pdx and supports monooxygenase reactions catalyzed by P450cam. The insights gained from these studies will be summarized in this review.

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