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