Skinner, SP, Follmer, AH, Ubbink, M et al. (3 more authors) (2021) Partial Opening of Cytochrome P450cam (CYP101A1) Is Driven by Allostery and Putidaredoxin Binding. Biochemistry. ISSN 0006-2960
Abstract
Cytochrome P450cam (CYP101A1) catalyzes the regio- and stereo-specific 5-exo-hydroxylation of camphor via a multistep catalytic cycle that involves two-electron transfer steps, with an absolute requirement that the second electron be donated by the ferrodoxin, putidaredoxin (Pdx). Whether P450cam, once camphor has bound to the active site and the substrate entry channel has closed, opens up upon Pdx binding, during the second electron transfer step, or it remains closed is still a matter of debate. A potential allosteric site for camphor binding has been identified and postulated to play a role in the binding of Pdx. Here, we have revisited paramagnetic NMR spectroscopy data and determined a heterogeneous ensemble of structures that explains the data, provides a complete representation of the P450cam/Pdx complex in solution, and reconciles alternative hypotheses. The allosteric camphor binding site is always present, and the conformational changes induced by camphor binding to this site facilitates Pdx binding. We also determined that the state to which Pdx binds comprises an ensemble of structures that have features of both the open and closed state. These results demonstrate that there is a finely balanced interaction between allosteric camphor binding and the binding of Pdx at high camphor concentrations.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2021 American Chemical Society. This is an author produced version of an article published in Biochemistry. Uploaded in accordance with the publisher's self-archiving policy. |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Biological Sciences (Leeds) > School of Molecular and Cellular Biology (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 28 Sep 2021 13:13 |
Last Modified: | 14 Sep 2022 00:15 |
Status: | Published online |
Publisher: | American Chemical Society (ACS) |
Identification Number: | 10.1021/acs.biochem.1c00406 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:178527 |