Fluorescent Labeling of the Leukocyte NADPH Oxidase Subunit p47phox: Evidence for Amphiphile-Induced Conformational Changes

1998 ◽  
Vol 360 (2) ◽  
pp. 165-172 ◽  
Author(s):  
Hee-Sae Park ◽  
Jeen-Woo Park
Keyword(s):  
Nadph Oxidase ◽  
Conformational Changes ◽  
Fluorescent Labeling ◽  
Oxidase Subunit

Phosphorylation of the leucocyte NADPH oxidase subunit p47phox by casein kinase 2: conformation-dependent phosphorylation and modulation of oxidase activity

2001 ◽  
Vol 358 (3) ◽  
pp. 783-790 ◽  
Author(s):  
Hee-Sae PARK ◽  
S. Min LEE ◽  
Jin Hyup LEE ◽  
Yun-Sook KIM ◽  
Young-Seuk BAE ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Conformational Changes ◽  
Oxidase Activity ◽  
Serine Threonine Kinase ◽  
Src Homology 3 ◽  
Oxidase Subunit ◽  
Putative Site ◽  
Src Homology ◽  
Membrane Bound ◽  
Eukaryotic Organisms

The leucocyte NADPH oxidase of neutrophils is a membrane-bound enzyme that catalyses the reduction of oxygen to O−2 at the expense of NADPH. The enzyme is dormant in resting neutrophils but becomes active when the cells are exposed to the appropriate stimuli. During oxidase activation, the highly basic cytosolic oxidase component p47phox becomes phosphorylated on several serines and migrates to the plasma membrane. Protein kinase CK2 is an essential serine/threonine kinase present in all eukaryotic organisms. The leucocyte NADPH oxidase subunit p47phox has several putative CK2 phosphorylation sites. In the present study, we report that CK2 is able to catalyse the phosphorylation of p47phoxin vitro. Phosphoamino acid analysis of phosphorylated p47phox by CK2 indicated that the phosphorylation occurs on serine residues. CNBr mapping and phosphorylation of peptides containing the putative site of CK2 indicated that the main phosphorylated residues are Ser-208 and Ser-283 in the Src homology 3 (SH3) domains, and Ser-348 in the C-terminal domain of p47phox. Dependence of phosphorylation on the conformation of p47phox is supported by the finding that p47phox undergoes better phosphorylation by CK2 in the presence of arachidonic acid, a known activator of NADPH oxidase which induces conformational changes in p47phox. In addition, 5,6-dichloro-1-β-o-ribofuranosyl benzimidazole, a CK2 inhibitor, potentiates formyl-Met-Leu-Phe-induced NADPH oxidase activity in DMSO-differentiated HL-60 cells. Taken together, we propose that CK2 is the p47phox kinase, and that phosphorylation of p47phox by CK2 regulates the deactivation of NADPH oxidase.

Effects of red grape juice polyphenols in NADPH oxidase subunit expression in human neutrophils and mononuclear blood cells

2009 ◽  
Vol 102 (8) ◽  
pp. 1125-1135 ◽  
Author(s):  
Alberto Dávalos ◽  
Gema de la Peña ◽  
Carolina C. Sánchez-Martín ◽  
M. Teresa Guerra ◽  
Begoña Bartolomé ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Red Wine ◽  
Grape Juice ◽  
Cell Types ◽  
Human Neutrophils ◽  
Superoxide Anion Production ◽  
Oxidase Subunit ◽  
Subunit Expression ◽  
Red Grape Juice ◽  
Red Grape

The NADPH oxidase enzyme system is the main source of superoxide anions in phagocytic and vascular cells. NADPH oxidase-dependent superoxide generation has been found to be abnormally enhanced in several chronic diseases. Evidence is accumulating that polyphenols may have the potential to improve cardiovascular health, although the mechanism is not fully established. Consumption of concentrated red grape juice, rich in polyphenols, has been recently shown to reduce NADPH oxidase activity in circulating neutrophils from human subjects. In the present work we studied whether red grape juice polyphenols affected NADPH oxidase subunit expression at the transcription level. For this, we used human neutrophils and mononuclear cells from peripheral blood, HL-60-derived neutrophils and the endothelial cell line EA.hy926.Superoxide production was measured with 2′7′-dichlorofluorescein diacetate or lucigenin, mRNA expression by real-time RT-PCR and protein expression by Western blot. Each experiment was performed at least three times. In all cell types tested, red grape juice, dealcoholised red wine and pure polyphenols decreased superoxide anion production. Red grape juice and dealcoholised red wine selectively reduced p47phox, p22phox and gp91phox expression at both mRNA and protein levels, without affecting the expression of p67phox. Pure polyphenols, particularly quercetin, also reduced NADPH oxidase subunit expression, especially p47phox, in all cell types tested. The present results showing that red grape juice polyphenols reduce superoxide anion production provide an alternative mechanism by which consumption of grape derivatives may account for a reduction of oxidative stress associated with cardiovascular and/or inflammatory diseases related to NADPH oxidase superoxide overproduction.

Resonance Raman Imaging of the NADPH Oxidase Subunit Cytochromeb558in Single Neutrophilic Granulocytes

2003 ◽  
Vol 125 (40) ◽  
pp. 12112-12113 ◽  
Author(s):  
Henk-Jan van Manen ◽  
Natallia Uzunbajakava ◽  
Robin van Bruggen ◽  
Dirk Roos ◽  
Cees Otto
Keyword(s):  
Nadph Oxidase ◽  
Resonance Raman ◽  
Raman Imaging ◽  
Neutrophilic Granulocytes ◽  
Oxidase Subunit

Anionic lipid-induced conformational changes in human phagocyte flavocytochrome b precede assembly and activation of the NADPH oxidase complex

2012 ◽  
Vol 521 (1-2) ◽  
pp. 24-31 ◽  
Author(s):  
Ross M. Taylor ◽  
Marcia H. Riesselman ◽  
Connie I. Lord ◽  
Jeannie M. Gripentrog ◽  
Algirdas J. Jesaitis
Keyword(s):  
Nadph Oxidase ◽  
Conformational Changes ◽  
Anionic Lipid ◽  
Nadph Oxidase Complex ◽  
Flavocytochrome B

Corrections - Conformational Changes in the Foot Protein of the Sarcoplasmic Reticulum Assessed by Site-Directed Fluorescent Labeling

Biochemistry ◽  
1992 ◽  
Vol 31 (20) ◽  
pp. 4922-4922
Author(s):  
J. Kang ◽  
A. Tarcsafalvi ◽  
A. Carlos ◽  
E. Fujimoto ◽  
Z. Shahrokh ◽  
...  
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Conformational Changes ◽  
Fluorescent Labeling

NADPH oxidase subunit NOXO1 is a target for emphysema treatment in COPD

2020 ◽  
Vol 2 (6) ◽  
pp. 532-546
Author(s):  
Michael Seimetz ◽  
Natascha Sommer ◽  
Mariola Bednorz ◽  
Oleg Pak ◽  
Christine Veith ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Oxidase Subunit
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