Direct Measurement by Laser Flash Photolysis of Intraprotein Electron Transfer in a Rat Neuronal Nitric Oxide Synthase

2007 ◽  
Vol 129 (17) ◽  
pp. 5621-5629 ◽  
Author(s):  
Changjian Feng ◽  
Gordon Tollin ◽  
James T. Hazzard ◽  
Nickolas J. Nahm ◽  
J. Guy Guillemette ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Electron Transfer ◽  
Nitric Oxide Synthase ◽  
Direct Measurement ◽  
Flash Photolysis ◽  
Laser Flash Photolysis ◽  
Neuronal Nitric Oxide Synthase ◽  
Laser Flash ◽  
Oxide Synthase

Dual Effects of the Substrate and Pterins on the Kinetics of CO Binding to Neuronal Nitric Oxide Synthase: A Laser Flash Photolysis Study

2005 ◽  
Vol 34 (6) ◽  
pp. 752-753
Author(s):  
Simona N. Bengea ◽  
Yasuyuki Araki ◽  
Osamu Ito ◽  
Jotaro Igarashi ◽  
Hirofumi Kurokawa ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Flash Photolysis ◽  
Laser Flash Photolysis ◽  
Neuronal Nitric Oxide Synthase ◽  
Laser Flash ◽  
Oxide Synthase ◽  
Kinetics Of

Haloperidol Inhibits Neuronal Nitric Oxide Synthase Activity by Preventing Electron Transfer

1996 ◽  
Vol 33 (2) ◽  
pp. 76-79 ◽  
Author(s):  
Kazuhiko Iwahashi ◽  
Hirohito Yoneyama ◽  
Taira Ohnishi ◽  
Kazuhiko Nakamura ◽  
Ryosuke Miyatake ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Electron Transfer ◽  
Nitric Oxide Synthase ◽  
Neuronal Nitric Oxide Synthase ◽  
Oxide Synthase ◽  
Nitric Oxide Synthase Activity

scholarly journals Stopped-flow kinetic studies of electron transfer in the reductase domain of neuronal nitric oxide synthase: re-evaluation of the kinetic mechanism reveals new enzyme intermediates and variation with cytochrome P450 reductase

2002 ◽  
Vol 367 (1) ◽  
pp. 19-30 ◽  
Author(s):  
Kirsty KNIGHT ◽  
Nigel S. SCRUTTON
Keyword(s):  
Nitric Oxide ◽  
Electron Transfer ◽  
Cytochrome P450 ◽  
Nitric Oxide Synthase ◽  
Neuronal Nitric Oxide Synthase ◽  
Stopped Flow ◽  
Cytochrome P450 Reductase ◽  
P450 Reductase ◽  
Oxide Synthase ◽  
Reductase Domain

The reduction by NADPH of the FAD and FMN redox centres in the isolated flavin reductase domain of calmodulin-bound rat neuronal nitric oxide synthase (nNOS) has been studied by anaerobic stopped-flow spectroscopy using absorption and fluorescence detection. We show by global analysis of time-dependent photodiode array spectra, single wavelength absorption and NADPH fluorescence studies, that at least four resolvable steps are observed in stopped-flow studies with NADPH and that flavin reduction is reversible. The first reductive step represents the rapid formation of an equilibrium between an NADPH-enzyme charge-transfer species and two-electron-reduced enzyme bound to NADP+. The second and third steps represent further reduction of the enzyme flavins and NADP+ release. The fourth step is attributed to the slow accumulation of an enzyme species that is inferred not to be relevant catalytically in steady-state reactions. Stopped-flow flavin fluorescence studies indicate the presence of slow kinetic phases, the timescales of which correspond to the slow phase observed in absorption and NADPH fluorescence transients. By analogy with stopped-flow studies of cytochrome P450 reductase, we attribute these slow fluorescence and absorption changes to enzyme disproportionation and/or conformational change. Unlike for the functionally related cytochrome P450 reductase, transfer of the first hydride equivalent from NADPH to nNOS reductase does not generate the flavin di-semiquinoid state. This indicates that internal electron transfer is relatively slow and is probably gated by NADP+ release. Release of calmodulin from the nNOS reductase does not affect the kinetics of inter-flavin electron transfer under stopped-flow conditions, although the observed rate of formation of the equilibrium between the NADPH-oxidized enzyme charge-transfer species and two-electron-reduced enzyme bound to NADP+ is modestly slower in calmodulin-depleted enzyme. Our studies indicate the need for significant re-interpretation of published kinetic data for electron transfer in the reductase domain of neuronal nitric oxide synthase.

Intraprotein Electron Transfer in a Two-Domain Construct of Neuronal Nitric Oxide Synthase:  The Output State in Nitric Oxide Formation†

Biochemistry ◽  
2006 ◽  
Vol 45 (20) ◽  
pp. 6354-6362 ◽  
Author(s):  
Changjian Feng ◽  
Gordon Tollin ◽  
Michael A. Holliday ◽  
Clayton Thomas ◽  
John C. Salerno ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Electron Transfer ◽  
Nitric Oxide Synthase ◽  
Neuronal Nitric Oxide Synthase ◽  
Oxide Formation ◽  
Output State ◽  
Nitric Oxide Formation ◽  
Oxide Synthase
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