Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength

Abstract Cytochrome P450 oxidoreductase (POR) is the primary electron donor in eukaryotic cytochrome P450 (CYP) containing systems. A wealth of ensemble biophysical studies of Cytochrome P450 oxidoreductase (POR) has reported a binary model of the conformational equilibrium directing its catalytic e...

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Autores principales:	Krutika Bavishi, Darui Li, Stine Eiersholt, Emma N. Hooley, Troels C. Petersen, Birger Lindberg Møller, Nikos S. Hatzakis, Tomas Laursen
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2018
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/8fa4bf61be47432a9c951c289cdc90d1
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spelling	oai:doaj.org-article:8fa4bf61be47432a9c951c289cdc90d12021-12-02T15:07:57ZDirect observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength10.1038/s41598-018-24922-x2045-2322https://doaj.org/article/8fa4bf61be47432a9c951c289cdc90d12018-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-018-24922-xhttps://doaj.org/toc/2045-2322Abstract Cytochrome P450 oxidoreductase (POR) is the primary electron donor in eukaryotic cytochrome P450 (CYP) containing systems. A wealth of ensemble biophysical studies of Cytochrome P450 oxidoreductase (POR) has reported a binary model of the conformational equilibrium directing its catalytic efficiency and biomolecular recognition. In this study, full length POR from the crop plant Sorghum bicolor was site-specifically labeled with Cy3 (donor) and Cy5 (acceptor) fluorophores and reconstituted in nanodiscs. Our single molecule fluorescence resonance energy transfer (smFRET) burst analyses of POR allowed the direct observation and quantification of at least three dominant conformational sub-populations, their distribution and occupancies. Moreover, the state occupancies were remodeled significantly by ionic strength and the nature of reconstitution environment, i.e. phospholipid bilayers (nanodiscs) composed of different lipid head group charges vs. detergent micelles. The existence of conformational heterogeneity in POR may mediate selective activation of multiple downstream electron acceptors and association in complexes in the ER membrane.Krutika BavishiDarui LiStine EiersholtEmma N. HooleyTroels C. PetersenBirger Lindberg MøllerNikos S. HatzakisTomas LaursenNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 8, Iss 1, Pp 1-9 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Krutika Bavishi Darui Li Stine Eiersholt Emma N. Hooley Troels C. Petersen Birger Lindberg Møller Nikos S. Hatzakis Tomas Laursen Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength
description	Abstract Cytochrome P450 oxidoreductase (POR) is the primary electron donor in eukaryotic cytochrome P450 (CYP) containing systems. A wealth of ensemble biophysical studies of Cytochrome P450 oxidoreductase (POR) has reported a binary model of the conformational equilibrium directing its catalytic efficiency and biomolecular recognition. In this study, full length POR from the crop plant Sorghum bicolor was site-specifically labeled with Cy3 (donor) and Cy5 (acceptor) fluorophores and reconstituted in nanodiscs. Our single molecule fluorescence resonance energy transfer (smFRET) burst analyses of POR allowed the direct observation and quantification of at least three dominant conformational sub-populations, their distribution and occupancies. Moreover, the state occupancies were remodeled significantly by ionic strength and the nature of reconstitution environment, i.e. phospholipid bilayers (nanodiscs) composed of different lipid head group charges vs. detergent micelles. The existence of conformational heterogeneity in POR may mediate selective activation of multiple downstream electron acceptors and association in complexes in the ER membrane.
format	article
author	Krutika Bavishi Darui Li Stine Eiersholt Emma N. Hooley Troels C. Petersen Birger Lindberg Møller Nikos S. Hatzakis Tomas Laursen
author_facet	Krutika Bavishi Darui Li Stine Eiersholt Emma N. Hooley Troels C. Petersen Birger Lindberg Møller Nikos S. Hatzakis Tomas Laursen
author_sort	Krutika Bavishi
title	Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength
title_short	Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength
title_full	Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength
title_fullStr	Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength
title_full_unstemmed	Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength
title_sort	direct observation of multiple conformational states in cytochrome p450 oxidoreductase and their modulation by membrane environment and ionic strength
publisher	Nature Portfolio
publishDate	2018
url	https://doaj.org/article/8fa4bf61be47432a9c951c289cdc90d1
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Direct observation of multiple conformational states in Cytochrome P450 oxidoreductase and their modulation by membrane environment and ionic strength

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