Biochemical and structural insights into the cytochrome P450 reductase from Candida tropicalis

Cytochrome P450 reductases (CPRs) are diflavin oxidoreductases that supply electrons to type II cytochrome P450 monooxygenases (CYPs). In addition, it can also reduce other proteins and molecules, including cytochrome c , ferricyanide, and different drugs. Although various CPRs have been functionall...

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Published in:	Scientific reports Vol. 9; no. 1; pp. 20088 - 11
Main Authors:	Ebrecht, Ana C., van der Bergh, Naadia, Harrison, Susan T. L., Smit, Martha S., Sewell, B. Trevor, Opperman, Diederik J.
Format:	Journal Article
Language:	English
Published:	London Nature Publishing Group UK 27-12-2019 Nature Publishing Group Nature Portfolio
Subjects:	631/45 631/45/535/1266 631/45/607/1168 631/61 Candida tropicalis Candida tropicalis - enzymology Conformation Crystallography, X-Ray Cytochrome Cytochrome c Cytochrome P450 Electron transfer Electron Transport Enzymes Ferricyanide Flavin mononucleotide Flavin-adenine dinucleotide Humanities and Social Sciences Hydrophobicity Intermediates Ionic strength multidisciplinary NAD NADP NADPH-ferrihemoprotein reductase NADPH-Ferrihemoprotein Reductase - chemistry NADPH-Ferrihemoprotein Reductase - metabolism Oxidation-Reduction Oxygen Protein Conformation Reactive oxygen species Science Science (multidisciplinary)
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Summary:	Cytochrome P450 reductases (CPRs) are diflavin oxidoreductases that supply electrons to type II cytochrome P450 monooxygenases (CYPs). In addition, it can also reduce other proteins and molecules, including cytochrome c , ferricyanide, and different drugs. Although various CPRs have been functionally and structurally characterized, the overall mechanism and its interaction with different redox acceptors remain elusive. One of the main problems regarding electron transfer between CPRs and CYPs is the so-called “uncoupling”, whereby NAD(P)H derived electrons are lost due to the reduced intermediates’ (FAD and FMN of CPR) interaction with molecular oxygen. Additionally, the decay of the iron-oxygen complex of the CYP can also contribute to loss of reducing equivalents during an unproductive reaction cycle. This phenomenon generates reactive oxygen species (ROS), leading to an inefficient reaction. Here, we present the study of the CPR from C andida tropicalis ( Ct CPR) lacking the hydrophobic N -terminal part (Δ2–22). The enzyme supports the reduction of cytochrome c and ferricyanide, with an estimated 30% uncoupling during the reactions with cytochrome c . The ROS produced was not influenced by different physicochemical conditions (ionic strength, pH, temperature). The X-ray structures of the enzyme were solved with and without its cofactor, NADPH. Both Ct CPR structures exhibited the closed conformation. Comparison with the different solved structures revealed an intricate ionic network responsible for the regulation of the open/closed movement of Ct CPR.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	2045-2322 2045-2322
DOI:	10.1038/s41598-019-56516-6