title: Fluorescence quenching studies of structure and dynamics in calmodulin-eNOS complexes.
abstract: Activation of endothelial nitric oxide synthase (eNOS) by calmodulin (CaM) facilitates formation of a sequence of conformational states that is not well understood. Fluorescence decays of fluorescently labeled CaM bound to eNOS reveal four distinct conformational states and single-molecule fluorescence trajectories show multiple fluorescence states with transitions between states occurring on time scales of milliseconds to seconds. A model is proposed relating fluorescence quenching states to enzyme conformations. Specifically, we propose that the most highly quenched state corresponds to CaM docked to an oxygenase domain of the enzyme. In single-molecule trajectories, this state occurs with time lags consistent with the oxygenase activity of the enzyme.
authors: Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA; Department of Chemistry, Northwestern College, Orange City, IA 51041, USA.
authors: David C
authors: DC
authors: Arnett
authors: Division of Molecular Biology and Biochemistry and Division of Cell Biology and Biophysics, University of Missouri at Kansas City, Kansas City, MO 64410, USA.
authors: Anthony
authors: A
authors: Persechini
authors: Division of Molecular Biology and Biochemistry and Division of Cell Biology and Biophysics, University of Missouri at Kansas City, Kansas City, MO 64410, USA.
authors: Quang-Kim
authors: QK
authors: Tran
authors: Division of Molecular Biology and Biochemistry and Division of Cell Biology and Biophysics, University of Missouri at Kansas City, Kansas City, MO 64410, USA.
authors: D J
authors: DJ
authors: Black
authors: Department of Chemistry, University of Kansas, Lawrence, KS 66045, USA. Electronic address: ckjohnson@ku.edu.
authors: Carey K
authors: CK
authors: Johnson
mesh_terms: D000818:Animals; D002147:Calmodulin / Q000737:chemistry*; D002417:Cattle; D036681:Fluorescence Recovery After Photobleaching; D005470:Fluorometry; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry*; D052250:Nitric Oxide Synthase Type III / Q000737:chemistry*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary
chemical_list: D002147:Calmodulin; D046912:Multiprotein Complexes; D052250:Nitric Oxide Synthase Type III
keywords: Calmodulin; Fluorescence decay; Förster resonance energy transfer; Nitric oxide synthase; Single-molecule fluorescence