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Endothelial Nitric-oxide Synthase

P. Chen, A. Tsai, V. Berka, K. Wu
Published 1996 · Medicine, Biology

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A baculovirus system was used to express the oxygenase and reductase domains of human endothelial nitric-oxide synthase (ecNOS) as distinct proteins. The oxygenase domain (residues 1–491) was expressed using a vector containing a His6 tag at the N terminus. The purified oxygenase domain had an apparent molecular mass of ∼54 kDa, and retained the ability to bind L-arginine and form the ferrous CO complex. The purified reductase domain (residues 492-1244) had an apparent molecular mass of ∼82 kDa and retained the ability to catalyze NADPH-dependent cytochrome c reduction, which was enhanced 10-fold by the presence of Ca2+/calmodulin. Both purified domains exhibited immunoreactivity to rabbit anti-ecNOS IgG. The NOS activity was successfully reconstituted by mixing the two domains. These results demonstrate for the first time that the two domains of ecNOS are catalytically intact and can be reconstituted in vitro.
This paper references
Evidence for a bidomain structure of constitutive cerebellar nitric oxide synthase.
E. Sheta (1994)
10.1016/0003-2697(76)90527-3
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding.
M. M. Bradford (1976)
10.1016/0006-2952(91)90581-O
Isoforms of nitric oxide synthase. Characterization and purification from different cell types.
U. Foerstermann (1991)
10.1016/0006-291X(91)91864-9
Inactivation of macrophage nitric oxide synthase activity by NG-methyl-L-arginine.
N. M. Olken (1991)
10.1073/PNAS.90.22.10769
Nitric oxide synthases reveal a role for calmodulin in controlling electron transfer.
H. Abu-Soud (1993)
10.1021/BI00036A003
Reconstitution of the second step in NO synthesis using the isolated oxygenase and reductase domains of macrophage NO synthase.
D. Ghosh (1995)
10.1016/0003-9861(89)90461-X
Novel exogenous heme-dependent expression of mammalian cytochrome P450 using baculovirus.
A. Asseffa (1989)
10.1126/SCIENCE.1373522
Cloning and characterization of inducible nitric oxide synthase from mouse macrophages.
Q. Xie (1992)
10.1021/BI00144A001
Nitric oxide synthase is a cytochrome P-450 type hemoprotein.
K. White (1992)
10.1038/227680A0
Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4
U. Laemmli (1970)
10.1021/BI00011A025
Prokaryotic expression of the heme- and flavin-binding domains of rat neuronal nitric oxide synthase as distinct polypeptides: identification of the heme-binding proximal thiolate ligand as cysteine-415.
K. McMillan (1995)
10.1073/PNAS.76.9.4350
Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications.
H. Towbin (1979)
10.1016/s0021-9258(19)50029-1
Reconstitution of the fatty acid hydroxylation function of cytochrome P-450BM-3 utilizing its individual recombinant hemo- and flavoprotein domains.
S. Boddupalli (1992)
10.1073/PNAS.92.18.8428
High-level expression of functional rat neuronal nitric oxide synthase in Escherichia coli.
L. Roman (1995)
10.1073/PNAS.89.23.11141
Cloned, expressed rat cerebellar nitric oxide synthase contains stoichiometric amounts of heme, which binds carbon monoxide.
K. McMillan (1992)
10.1073/PNAS.88.23.10480
Purification and characterization of particulate endothelium-derived relaxing factor synthase from cultured and native bovine aortic endothelial cells.
J. Pollock (1991)
10.1016/s0021-9258(19)36718-3
Spectral characterization of brain and macrophage nitric oxide synthases. Cytochrome P-450-like hemeproteins that contain a flavin semiquinone radical.
D. Stuehr (1992)
10.1006/BBRC.1995.1659
Modular structure of neuronal nitric oxide synthase: localization of the arginine binding site and modulation by pterin.
J. S. Nishimura (1995)
10.1038/351714A0
Cloned and expressed nitric oxide synthase structurally resembles cytochrome P-450 reductase
D. Bredt (1991)
10.1016/s0021-9258(18)38112-2
N omega-hydroxy-L-arginine is an intermediate in the biosynthesis of nitric oxide from L-arginine.
D. Stuehr (1991)
10.1042/BJ2880015
Brain nitric oxide synthase is a haemoprotein.
P. Klatt (1992)
Cysteine 184 of endothelial nitric oxide synthase is involved in heme coordination and catalytic activity.
P. Chen (1994)
10.1006/ABBI.1995.1356
N-nitro-L-arginine and N-monomethyl-L-arginine exhibit a different pattern of inactivation toward the three nitric oxide synthases.
D. Reif (1995)
10.1021/BI00003A013
Macrophage NO synthase: characterization of isolated oxygenase and reductase domains reveals a head-to-head subunit interaction.
D. Ghosh (1995)
10.1096/fasebj.6.12.1381691
Nitric oxide as a secretory product of mammalian cells
C. Nathan (1992)
10.1016/s0021-9258(19)36901-7
Macrophage nitric oxide synthase subunits. Purification, characterization, and role of prosthetic groups and substrate in regulating their association into a dimeric enzyme.
K. J. Baek (1993)
10.1016/s0021-9258(18)54421-5
Purification of the inducible murine macrophage nitric oxide synthase. Identification as a flavoprotein.
J. Hevel (1991)
10.1021/BI00146A019
Macrophage nitric oxide synthase: relationship between enzyme-bound tetrahydrobiopterin and synthase activity.
J. Hevel (1992)
10.1021/BI00127A028
Ca2+/calmodulin-dependent NO synthase type I: a biopteroflavoprotein with Ca2+/calmodulin-independent diaphorase and reductase activities.
H. Schmidt (1992)
10.1006/BBRC.1995.2579
Cysteine 99 of endothelial nitric oxide synthase (NOS-III) is critical for tetrahydrobiopterin-dependent NOS-III stability and activity.
P. Chen (1995)
10.1006/BBRC.1993.1380
Particular ability of liver P450s3A to catalyze the oxidation of N omega-hydroxyarginine to citrulline and nitrogen oxides and occurrence in no synthases of a sequence very similar to the heme-binding sequence in P450s.
J. P. Renaud (1993)
10.1016/s0021-9258(18)48296-8
Identification and characterization of two functional domains in cytochrome P-450BM-3, a catalytically self-sufficient monooxygenase induced by barbiturates in Bacillus megaterium.
L. Narhi (1987)
10.1073/PNAS.87.2.682
Isolation of nitric oxide synthetase, a calmodulin-requiring enzyme.
D. Bredt (1990)
10.1021/BI00089A001
Optical difference spectrophotometry as a probe of rat brain nitric oxide synthase heme-substrate interaction.
K. McMillan (1993)
10.1073/PNAS.88.17.7773
Purification and characterization of the cytokine-induced macrophage nitric oxide synthase: an FAD- and FMN-containing flavoprotein.
D. Stuehr (1991)
10.1073/PNAS.86.22.9030
Nitric oxide mediates glutamate-linked enhancement of cGMP levels in the cerebellum.
D. Bredt (1989)
10.1016/s0021-9258(18)31375-9
Nitric oxide synthase structure and mechanism.
M. Marletta (1993)
10.1073/pnas.74.12.5463
DNA sequencing with chain-terminating inhibitors.
F. Sanger (1977)
10.1016/s0021-9258(19)49920-1
Ca2+/calmodulin-dependent cytochrome c reductase activity of brain nitric oxide synthase.
P. Klatt (1992)



This paper is referenced by
10.1074/jbc.274.35.24694
Geminate Recombination of Nitric Oxide to Endothelial Nitric-oxide Synthase and Mechanistic Implications*
M. Negrerie (1999)
10.1074/jbc.M003935200
Mutations at Lysine 525 of Inducible Nitric-oxide Synthase Affect Its Ca2+-independent Activity*
S. Lee (2000)
10.1074/JBC.M105691200
Mechanistic Studies with Potent and Selective Inducible Nitric-oxide Synthase Dimerization Inhibitors*
E. Blasko (2002)
10.5897/AJB12.2886
Xanthine oxidase activity during transition period and its association with occurrence of postpartum infections in Murrah buffalo ( Bubalus bubalis )
B. Mili (2013)
10.1093/emboj/18.22.6260
Inducible nitric oxide synthase: role of the N‐terminal β‐hairpin hook and pterin‐binding segment in dimerization and tetrahydrobiopterin interaction
D. Ghosh (1999)
10.18388/abp.2002_3844
Polymorphism in intron 23 of the endothelial nitric oxide synthase gene (NOS3) is not associated with hypertension.
Natalia Derebecka (2002)
10.1074/jbc.M100687200
Calmodulin Activates Intersubunit Electron Transfer in the Neuronal Nitric-oxide Synthase Dimer*
K. Panda (2001)
10.1074/JBC.M209606200
Redox Properties of Human Endothelial Nitric-oxide Synthase Oxygenase and Reductase Domains Purified from Yeast Expression System*
M. Du (2003)
10.1006/BBRC.1998.8893
Chiral recognition at the heme active site of nitric oxide synthase is markedly enhanced by L-arginine and 5,6,7,8-tetrahydrobiopterin.
K. Nakano (1998)
10.1042/bj3570593
Nitric oxide synthases: structure, function and inhibition.
W. K. Alderton (2001)
10.1074/jbc.M802914200
Differences in a Conformational Equilibrium Distinguish Catalysis by the Endothelial and Neuronal Nitric-oxide Synthase Flavoproteins*
Robielyn P. Ilagan (2008)
10.1016/S0272-6386(97)90275-4
Nitric oxide in renal health and disease.
B. Kone (1997)
10.1016/J.CPLETT.2017.04.081
Ultrafast relaxation dynamics of nitric oxide synthase studied by visible broadband transient absorption spectroscopy
Chih Chang Hung (2017)
10.1042/BJ20100960
Reduction of N(ω)-hydroxy-L-arginine by the mitochondrial amidoxime reducing component (mARC).
J. Kotthaus (2011)
10.1007/978-3-642-57077-3_3
Enzymology of Nitric Oxide Synthases
D. Stuehr (2000)
10.1016/S0014-5793(00)01616-1
Catalase activity of oxygenase domain of rat neuronal nitric oxide synthase. Evidence for product formation from L‐arginine
S. Adhikari (2000)
10.1074/JBC.M403892200
Differential Activation of Nitric-oxide Synthase Isozymes by Calmodulin-Troponin C Chimeras*
E. Newman (2004)
10.1016/S0076-6879(99)01074-5
Scintillation proximity assay to measure nitroarginine and tetrahydrobiopterin binding to heme domain of neuronal nitric oxide synthase.
W. K. Alderton (1999)
10.1016/S0304-4165(99)00193-2
Calmodulin activates intramolecular electron transfer between the two flavins of neuronal nitric oxide synthase flavin domain.
H. Matsuda (1999)
10.1074/jbc.271.52.33293
Spatial Relationship between L-Arginine and Heme Binding Sites of Endothelial Nitric-oxide Synthase*
V. Berka (1996)
10.22037/JPS.V3I1.2915
The analysis of endothelial nitric oxide synthase gene polymorphism in intron 4 with hypertension disease
Leila Zarmakhi (2012)
10.1039/B506355H
Exploring the redox reactions between heme and tetrahydrobiopterin in the nitric oxide synthases.
D. Stuehr (2005)
10.1007/978-3-7091-6351-1_1
Nitric oxide: physiological roles, biosynthesis and medical uses.
D. R. Adams (1999)
10.1146/ANNUREV.PHARMTOX.37.1.339
Structure-function aspects in the nitric oxide synthases.
D. Stuehr (1997)
10.1074/jbc.272.28.17349
The Ferrous-dioxy Complex of Neuronal Nitric Oxide Synthase
H. Abu-Soud (1997)
10.1074/JBC.M108657200
Nitric Oxide (NO) Traffic in Endothelial NO Synthase
A. Slama-Schwok (2002)
10.1016/S0003-9861(02)00729-4
Differential effects of mutations in human endothelial nitric oxide synthase at residues Tyr-357 and Arg-365 on L-arginine hydroxylation and GN-hydroxy-L-arginine oxidation.
P. Chen (2003)
10.1371/journal.pone.0039851
Lobe-Specific Calcium Binding in Calmodulin Regulates Endothelial Nitric Oxide Synthase Activation
Pei-Rung Wu (2012)
10.1016/J.YJMCC.2004.05.004
Arginase pathway in human endothelial cells in pathophysiological conditions.
T. Bachetti (2004)
10.1074/jbc.273.35.22267
Interaction between Caveolin-1 and the Reductase Domain of Endothelial Nitric-oxide Synthase
S. Ghosh (1998)
10.1093/cvr/cvt267
A novel truncated form of eNOS associates with altered vascular function.
E. Galluccio (2014)
10.1074/JBC.275.5.3201
The Ferrous Dioxygen Complex of the Oxygenase Domain of Neuronal Nitric-oxide Synthase*
M. Couture (2000)
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