Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:1.6.3.1 (NADPH oxidase)
 11,281 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The superoxide-generating NADPH oxidase system in phagocytes consists of at least membrane-associated cytochrome b558 and three cytosolic components named SOCI/NCF-3/sigma 1/C1, SOCII/NCF-1/p47-phox, and SO-CIII/NCF-2/p67-phox. p47-phox and p67-phox were isolated, and their primary structures were determined, but SOCI has not been well characterized. In the present study, we first purified SOCI to homogeneity from the cytosol fraction of the differentiated HL-60 cells. The purified SOCI was a small GTP-binding protein (G protein) with a M(r) of about 22,000. The guanosine 5'-(3-O-thio)triphosphate-bound form, but not the GDP-bound form, of this small G protein showed the SOCI activity. The partial amino acid sequence of SOCI thus far determined was identical to the amino acid sequence deduced from the cDNA encoding rac2 p21. None of the purified small G proteins, including Ki-ras p21, smg p21B/rap1B p21, rhoA p21, and rac1 p21, showed the SOCI activity. These results indicate that SOCI is a small G protein very similar, if not identical, to rac2 p21. The GDP/GTP exchange reaction of SOCI was stimulated and inhibited by stimulatory and inhibitory GDP/GTP exchange proteins for small G proteins, named smg GDS and rho GDI, respectively. The NADPH oxidase activity was also stimulated and inhibited by smg GDS and rho GDI, respectively. These results indicate that the superoxide-generating NADPH oxidase system is regulated by both smg GDS and rho GDI through rac2 p21 or the rac2-related small G protein in phagocytes.
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PMID:Regulation of the superoxide-generating NADPH oxidase by a small GTP-binding protein and its stimulatory and inhibitory GDP/GTP exchange proteins. 131 93

During the past 5 years, the discovery of cell-free superoxide generation system (Bromberg Y, Pick E: Cell Immunol 88:213-221, 1984) has been revolutionized our understanding of phagocyte superoxide generation. Using cell-free system, it was clarified that NADPH oxidase for superoxide generation was comprised of components present in both the plasma membrane as well as in the cytosol. This oxidase could be kept inactive by keeping its components separated from each other within the cell and then quickly bringing them together in the plasma membrane upon activation. We analyzed cytosol components with column method, and clarified that 3 neutrophil cytosol factors (NCF-1/-2/-3) was necessary for reconstitute the cytosol activity which was missing in an autosomal recessive type of Chronic Granulomatous Disease (CGD) patients (Nunoi H, et al:Science 242:1298-1301, 1988). NCF-1/-2 were analyzed with B1 antibody and found their molecular weight as 47 and 67 kilodalton respectively. One of autosomal CGD patients was missing NCF-67k and the others were missing NCF-47k. NCF-47k and -67k were cloned with this antibody and sequenced and expressed as recombinant NCF-47k/-67k using baculovirus/insect cell system. Using these recombinants, we are trying to purify NCF-3 which is reported as small G protein in these days. Using monoclonal antibodies against these recombinants, we analyzed tissues with immunohistochemical methods and are trying to classify the type of CGD patients in Japan. In summary, at least five oxidase components now have been identified (alpha and beta chain of cytochrome b558 (gp91-phox, p22-phox) and NCF-47k/-67k/-3 (p47-phox/p67-phox/delta-1 or SOCI)).
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PMID:[Molecular bases of chronic granulomatous disease--analysis of the involvement of cytosol factors for NADPH oxidase]. 131 71

Phagocytic leukocytes contain an activatable NADPH:O2 oxidoreductase. Components of this enzyme system include cytochrome b558, and three soluble oxidase components (SOC I, SOC II, and SOC III) found in the cytosol of resting cells. Previously, we found that SOC II copurifies with, and is probably identical to, a 47-kDa substrate of protein kinase C. In the present study we investigated the change in location of several of these oxidase components after activation of intact neutrophils with phorbol myristate acetate (PMA) and separation of subcellular fraction on sucrose density gradients. On Western blots with fractions of resting cells, the alpha subunit of cytochrome b558 was detected with a monoclonal antibody as a doublet of Mr 22,000 and 24,000 in the specific granules and as a single band of Mr 24,000 in the plasma membrane. PMA induced an increase of cytochrome b558 in the plasma membrane, including the Mr 22,000 band. PMA also induced translocation of the 47-kDa protein from the cytosol to the membrane fraction, as revealed by in vitro phosphorylation experiments. When NADPH oxidase activity was determined in a cell-free system in the presence of sodium dodecyl sulfate and GTP with plasma membranes from resting cells, cytosol from PMA-treated cells was deficient compared with cytosol from resting cells. This deficiency could be partially restored by the addition of SOC I. Concomitantly, SOC I activity appeared in the plasma membranes of PMA-treated cells. These studies support the hypothesis that PMA stimulation of neutrophils results in assembly of oxidase components from the cytosol and the specific granules in the plasma membrane with subsequent expression of NADPH oxidase activity.
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PMID:Assembly and activation of the NADPH:O2 oxidoreductase in human neutrophils after stimulation with phorbol myristate acetate. 215 19

Neutrophil NADPH:O2 oxidoreductase activity, essential in the killing of bacteria by neutrophils, can be elicited in a cell-free system that requires plasma membranes, cytosol and sodium dodecyl sulfate. In addition, GTP or its nonhydrolyzable analog guanosine 5'-3-O-(thio)triphosphate (GTP gamma S) enhances NADPH oxidase activity. We investigated the mechanism of this effect of GTP gamma S in the cell-free system. Cytosol from human neutrophils was separated in three different soluble oxidase components (SOC I, SOC II, and SOC III). Previously we (Bolscher, B. G. J. M., Van Zwieten, R., Kramer, I. J. M., Weening, R. S., Verhoeven, A. J., and Roos, D. (1989) J. Clin. Invest. 83, 757-763) reported that the cytosol contains two components which act synergistically. We now report that one component (previously labeled SOC II) contains two different components that can be separated by ion exchange chromatography. Immunoblotting with antiserum B-1 (Volpp, B. D., Nauseef, W. M., and Clark, R. A. (1988) Science 242, 1295-1297), directed against a cytosolic complex capable of activating latent membranes in the cell-free system, showed a 47-kDa protein in SOC II and a 67-kDa protein in SOC III. SOC II also contains the 47-kDa phosphoprotein, which indicates that this phosphoprotein and the protein recognized by the antiserum are identical. Low rates of NADPH-dependent O2 consumption can be elicited by SOC II and SOC III in the absence of SOC I. This activity is independent of GTP gamma S. Addition of SOC I increases this activity 3-4-fold, only when GTP gamma S is present. Plasma membranes, incubated with SOC I plus GTP gamma S and re-isolated, showed a similar 3-4-fold enhanced O2 consumption with SOC II and SOC III. The GTP gamma S effect is exerted primarily at the level of the plasma membrane. The concentration of GTP gamma S that causes a half-maximal stimulation was 0.4 mu M. It is concluded that SOC I is a functional component of the NADPH oxidase.
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PMID:The activity of one soluble component of the cell-free NADPH:O2 oxidoreductase of human neutrophils depends on guanosine 5'-O-(3-thio)triphosphate. 220 87

Hypoxic exposure causes pulmonary vasoconstriction, which serves as a critical physiologic process that ensures regional alveolar ventilation and pulmonary perfusion in the lungs, but may become an essential pathologic factor leading to pulmonary hypertension. Although the molecular mechanisms underlying hypoxic pulmonary vasoconstriction and associated pulmonary hypertension are uncertain, increasing evidence indicates that hypoxia can result in a significant increase in intracellular reactive oxygen species concentration ([ROS](i)) through the mitochondrial electron-transport chain in pulmonary artery smooth muscle cells (PASMCs). The increased mitochondrial ROS subsequently activate protein kinase C-epsilon (PKCepsilon) and NADPH oxidase (Nox), providing positive mechanisms that further increase [ROS](i). ROS may directly cause extracellular Ca(2+) influx by inhibiting voltage-dependent K(+) (K(V)) channels and opening of store-operated Ca(2+) (SOC) channels, as well as intracellular Ca(2+) release by activating ryanodine receptors (RyRs), leading to an increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) and associated contraction. In concert with ROS, PKCepsilon may also affect K(V) channels, SOC channels, and RyRs, contributing to hypoxic Ca(2+) and contractile responses in PASMCs.
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PMID:ROS-dependent signaling mechanisms for hypoxic Ca(2+) responses in pulmonary artery myocytes. 1976 82