Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

p67phox is an essential part of the NADPH oxidase, a multiprotein enzyme complex that produces superoxide ions in response to microbial infection. Binding of the small GTPase Rac to p67phox is a key step in the assembly of the active enzyme complex. The structure of Rac.GTP bound to the N-terminal TPR (tetratricopeptide repeat) domain of p67phox reveals a novel mode of Rho family/effector interaction and explains the basis of GTPase specificity. Complex formation is largely mediated by an insertion between two TPR motifs, suggesting an unsuspected versatility of TPR domains in target recognition and in their more general role as scaffolds for the assembly of multiprotein complexes.
Mol Cell 2000 Oct
PMID:Structure of the TPR domain of p67phox in complex with Rac.GTP. 1109 Jun 27

It is established that agmatine, an endogenously formed decarboxylated arginine, is a weak competitive inhibitor of neuronal nitric-oxide synthase (nNOS) with an apparent Ki value of 660 microM [Biochem J 316:247-249, 1996]. Although agmatine is known to bind to alpha-adrenergic and imidazoline receptors, it has been suggested that some of the pharmacological actions of agmatine, such as the prevention of morphine tolerance, may be due to the inhibition of nNOS. In the current study, we have discovered that agmatine, at concentrations much lower than the reported Ki value, leads to a time-, concentration-, NADPH-, and calmodulin-dependent irreversible inactivation of nNOS. The kinetics of inactivation could be described by an apparent dissociation constant for the initial reversible complex (Ki) and a pseudo first-order inactivation constant (k(inact)) of 29 microM and 0.01 min(-1), respectively. As determined by high-performance liquid chromatography analysis, the mechanism of inactivation involves alteration of the prosthetic heme moiety of nNOS, in part to protein-bound products. Moreover, we discovered that agmatine causes a 3-fold increase in the NADPH oxidase activity of nNOS leading to the production of H2O2 and is a likely cause for the inactivation of the enzyme. Both the inactivation of nNOS and the oxidative stress produced should now be considered in the pharmacological actions of agmatine as well as provide insight into the potential biological effects of endogenously formed agmatine.
Mol Pharmacol 2001 Jan
PMID:Agmatine enhances the NADPH oxidase activity of neuronal NO synthase and leads to oxidative inactivation of the enzyme. 1112 20

Low-level arsenite treatment of porcine aortic endothelial cells (PAEC) stimulated superoxide accumulation that was attenuated by inhibitors of NAD(P)H oxidase. To demonstrate whether arsenite stimulated NADPH oxidase, intact PAEC were treated with arsenite for up to 2 h and membrane fractions were prepared to measure NADPH oxidase activity. Arsenite (5 microM) stimulated a twofold increase in activity by 1 h, which was inhibited by the oxidase inhibitor diphenyleneiodonium chloride. Direct treatment of isolated membranes with arsenite had no effect. Analysis of NADPH oxidase components revealed that p67(phox) localized exclusively to membranes of both control and treated cells. In contrast, cytosolic Rac1 translocated to the membrane fractions of cells treated with arsenite or angiotensin II but not with tumor necrosis factor. Immunodepletion of p67(phox) blocked oxidase activity stimulated by all three compounds. However, depleting Rac1 inhibited responses only to arsenite and angiotensin II. These data demonstrate that stimulus-specific activation of NADPH oxidase in endothelial cells was the source of reactive oxygen in endothelial cells after noncytotoxic arsenite exposure.
Am J Physiol Lung Cell Mol Physiol 2001 Mar
PMID:Arsenite stimulates plasma membrane NADPH oxidase in vascular endothelial cells. 1115 27

Endotoxin (lipopolysaccharide [LPS]) is known to induce the production of tumor necrosis factor (TNF)-alpha and the induction of manganese superoxide dismutase (MnSOD). We have recently demonstrated that induction of TNF-alpha and MnSOD by LPS is mediated through different signal transduction pathways. In the current study, we investigated the role of reactive oxygen species (ROS) in the induction of TNF-alpha and MnSOD messenger RNAs (mRNAs) in human monocytes. Hypoxia (1% O2) inhibited the production of superoxide (O2-) and the induction of MnSOD, but not TNF-alpha, mRNA. Diphenylene iodonium (DPI), a potent inhibitor of the reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, had no effect on LPS induction of MnSOD mRNA, whereas it markedly inhibited LPS-induced O2- production. Neither hypoxia nor DPI had any effect on LPS activation of nuclear factor (NF)-kappaB. These results suggest that (1) ROS is important in the induction of MnSOD, but not TNF-alpha, mRNA by LPS, (2) ROS from sources other than NADPH oxidase is involved in LPS induction of MnSOD mRNA, and (3) ROS-mediated LPS induction of MnSOD mRNA is independent of NF-kappaB activation.
Am J Respir Cell Mol Biol 2001 Feb
PMID:Differential induction of TNF-alpha and MnSOD by endotoxin: role of reactive oxygen species and NADPH oxidase. 1115 50

It has been suggested that human spermatozoa contain an NADPH oxidase that could generate reactive oxygen species involved in signalling pathways to promote fertility. The proposal depends on observations that the addition of NADPH to purified human spermatozoa stimulates chemiluminescence by the superoxide (O2-) probe, lucigenin. We confirmed these observations, but demonstrated that lucigenin increases NADPH consumption by spermatozoa and stimulates artefactual O2- production via a diphenyleneiodonium (DPI) sensitive flavoprotein. In the absence of cytochrome c, DPI-inhibitable NADPH oxidation by permeabilized spermatozoa was 8 times too small to account for the rate of NADPH-stimulated cytochrome c reduction. Thus NADPH can directly reduce cytochrome c by a flavoprotein dependent mechanism making this O2- assay also unreliable in sperm suspensions. We were unable to observe O2- production by 40 x 10(6) spermatozoa/ml using electron paramagnetic resonance spectroscopy but could identify O(2)(-) generation from 2000 4beta-phorbol-12-myristate-13-actetate (PMA)-stimulated leukocytes. Using spectrophotometry, we did not detect the reduced cytochrome b(558) component of the neutrophil NADPH oxidase in human spermatozoa. No hydrogen peroxide generation was observed using a sensitive Amplex Red assay. We conclude that human spermatozoa do not possess significant NADPH oxidase activity and that the mechanism by which NADPH promotes capacitation must be re-evaluated.
Mol Hum Reprod 2001 Mar
PMID:A critical investigation of NADPH oxidase activity in human spermatozoa. 1122 43

Hypoxic pulmonary vasoconstriction (HPV) matches lung perfusion with ventilation but may also result in chronic pulmonary hypertension. It has not been clarified whether acute HPV and the response to prolonged alveolar hypoxia are triggered by identical mechanisms. We characterized the vascular response to sustained hypoxic ventilation (3% O(2) for 120-180 min) in isolated rabbit lungs. Hypoxia provoked a biphasic increase in pulmonary arterial pressure (PAP). Persistent PAP elevation was observed after termination of hypoxia. Total blockage of lung nitric oxide (NO) formation by N(G)-monomethyl-L-arginine caused a two- to threefold amplification of acute HPV, the sustained pressor response, and the loss of posthypoxic relaxation. This amplification was only moderate when NO formation was partially blocked by the inducible NO synthase inhibitor S-methylisothiourea. The superoxide scavenger nitro blue tetrazolium and the superoxide dismutase inhibitor triethylenetetramine reduced the initial vasoconstrictor response, the prolonged PAP increase, and the loss of posthypoxic vasorelaxation to a similar extent. The NAD(P)H oxidase inhibitor diphenyleneiodonium nearly fully blocked the late vascular responses to hypoxia in a dose that effected a decrease to half of the acute HPV. In conclusion, as similarly suggested for acute HPV, lung NO synthesis and the superoxide-hydrogen peroxide axis appear to be implicated in the prolonged pressor response and the posthypoxic loss of vasorelaxation in perfused rabbit lungs undergoing 2-3 h of hypoxic ventilation.
Am J Physiol Lung Cell Mol Physiol 2001 Apr
PMID:NO and reactive oxygen species are involved in biphasic hypoxic vasoconstriction of isolated rabbit lungs. 1123 3

The role of oxidants in the mechanism of tumor promotion by peroxisome proliferators remains controversial. The idea that induction of acyl-coenzyme A oxidase leads to increased production of H(2)O(2), which damages DNA, seems unlikely; still, free radicals might be important in signaling in specialized cell types such as Kupffer cells, which produce mitogens. Because hard evidence for increased oxidant production in vivo after treatment with peroxisome proliferators is lacking, the spin-trapping technique and electron spin resonance spectroscopy were used. Rats were given di(2-ethylhexyl) phthalate (DEHP) acutely. The spin trapping agent alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone was also given and bile samples were collected for 4 h. Under these conditions, the intensity of the six-line radical adduct signal increased to a maximum value of 2.5-fold 2 h after administration of DEHP, before peroxisomal oxidases were induced. Furthermore, DEHP given with [(13)C(2)]dimethyl sulfoxide produced a 12-line electron spin resonance spectrum, providing evidence that DEHP stimulates (*)OH radical formation in vivo. Furthermore, when rats were pretreated with dietary glycine, which inactivates Kupffer cells, DEHP did not increase radical signals. Moreover, similar treatments were performed in knockout mice deficient in NADPH oxidase (p47(phox) subunit). Importantly, DEHP increased oxidant production in wild-type but not in NADPH oxidase-deficient mice. These data provide evidence for the hypothesis that the molecular source of free radicals induced by peroxisome proliferators is NADPH oxidase in Kupffer cells. On the contrary, radical adduct formation was not affected in peroxisome proliferator-activated receptor alpha knockout mice. These observations represent the first direct, in vivo evidence that phthalates increase free radicals in liver before peroxisomal oxidases are induced.
Mol Pharmacol 2001 Apr
PMID:Phthalates rapidly increase production of reactive oxygen species in vivo: role of Kupffer cells. 1125 18

The oxidative burst has been suggested to be a primary event responsible for triggering the cascade of defense responses in various plant species against infection with avirulent pathogens or pathogen-derived elicitors. The molecular mechanisms of rapid production of active oxygen species (AOS), however, are not well known. We isolated homologs of gp91 phox, a plasma membrane protein of the neutrophil NADPH oxidase, from a potato cDNA library. Molecular cloning of the cDNA showed that there are two isogenes, designated StrbohA and StrbohB, respectively. The RNA gel blot analyses showed that StrbohA was constitutively expressed at a low level, whereas StrbohB was induced by hyphal wall components (HWC elicitor) from Phytophthora infestans in potato tubers. Treatment of potato tubers with HWC elicitor caused a rapid but weak transient accumulation of H2O2 (phase I), followed by a massive oxidative burst 6 to 9 h after treatment (phase II). Diphenylene iodonium (DPI), an inhibitor of the neutrophil NADPH oxidase, blocked both bursts, whereas pretreatment of the protein synthesis inhibitor cycloheximide with the tuber abolished only the second burst. These results suggest that the expression of StrbohA and StrbohB contributes to phase I and II bursts, respectively. The same is true for arachidonic acid, a lipid component of P. infestans-stimulated biphasic oxidative burst, whereas an endogenous signaling molecule, salicylic acid, only induced a weak phase II burst. Both molecules induced the StrbohB expression, which is in agreement with the second burst. To characterize the signal transduction pathway leading to the oxidative burst, we examined the role of protein phosphorylation in HWC-stimulated StrbohB gene expression. K252a and staurosporine, two protein kinase inhibitors, blocked the transcript accumulation. Two inhibitors of extracellular Ca2+ movement, however, did not abolish the transcript accumulation of StrbohB, suggesting that certain calcium-independent protein kinases are involved in the process of StrbohB gene expression. Additionally, we examined a causal relationship between the oxidative burst and expression of defense genes induced by the HWC elicitor. The transcript accumulation of genes related to sesquiterpenoid phytoalexin synthesis (lubimin and rishitin) and phenylpropanoid pathway was inhibited slightly by the DPI treatment, suggesting that the oxidative burst is not essential to activate these genes. Interestingly, the concomitant presence of DPI with the elicitor resulted in an increase in lubimin accumulation and a decrease in rishitin accumulation. Because it is known that lubimin is metabolized into rishitin via oxylubimin, we propose that AOS mediates the synthesis of rishitin from lubimin.
Mol Plant Microbe Interact 2001 Jun
PMID:Induction of plant gp91 phox homolog by fungal cell wall, arachidonic acid, and salicylic acid in potato. 1138 68

Coronary microvascular endothelial cells exert (patho)physiological effects on the function of cardiac myocytes, which may be studied experimentally using pure cell populations. As an essential pre-requisite to the investigation of cells from gene-modified mice, we studied the phenotypic properties of coronary microvascular endothelial cells isolated from normal mice, and biochemically characterized the superoxide production by these cells. Microvascular endothelial cells were isolated from devitalized mouse ventricular tissue after sequential digestion with collagenase, trypsin and DNase. Coronary microvascular endothelial cells were separated from cardiac myocytes and other cells by differential centrifugation, plating and culture. Mouse coronary microvascular endothelial cells showed an irregular "cobblestone" morphology at confluence, were >98% positive for CD31 by FACS analysis, and were also positive for VE-cadherin and endothelial-type nitric oxide synthase (eNOS) by confocal microscopy. The cells took up fluorescently labelled, acetylated low-density lipoprotein, but were negative for a alpha -smooth muscle actin, desmin and cytokeratin. Unlike human endothelial cells, mouse coronary microvascular endothelial cells only weakly expressed von Willebrand factor. Immunoblotting showed that the mouse cells expressed components of a phagocyte-type NADPH oxidase. They exhibited NADPH-dependent O(2)(-)-generating activity, which was increased by angiotensin II but completely inhibited by diphenyleneiodonium. Thus, mouse coronary microvascular endothelial cells express both eNOS and NADPH oxidase, interactions between which may play a role in endothelial cell pathophysiology.
J Mol Cell Cardiol 2001 Jun
PMID:Phenotypic properties and characteristics of superoxide production by mouse coronary microvascular endothelial cells. 1144 17

We examined the subcellular localizations of NAD(P)H oxidase, a reactive oxygen species (ROS)-producing enzyme, in fetal membrane chorion laeve trophoblasts from preterm or term pregnant women with or without chorioamnionitis (CAM). Ultrastructural enzyme histochemistry for NAD(P)H oxidase was used. In fetal membranes without CAM, approximately one quarter of the chorion laeve trophoblasts (25.6%) showed NAD(P)H oxidase activity on their surface plasma membranes and microvillous membranes. In mild CAM, the proportion of these NAD(P)H oxidase-positive cells significantly increased, reaching about half (51.0%). Enzyme activity appeared on the plasma and microvillous membranes and also on both phagosomal membranes and intracellular vesico-tubular structures. Appearance of NAD(P)H oxidase on surface plasma membranes, phagosomal membranes, and vesico-tubular structures is strong cytochemical evidence of phagocytic cell activation. These observations indicate that chorion laeve trophoblasts possess NAD(P)H oxidase activity, and therefore that fetal membranes themselves have ROS-generating capacity. Further, in fetal membrane inflammation, chorion laeve trophoblasts exhibited enzyme distribution characteristic of activated professional phagocytes. Similar to phagocytes infiltrating to the intrauterine environment, chorion laeve trophoblast NAD(P)H oxidase may play a role both in the defence of chorioamnion against infection and in the pathogenesis or pathophysiology of CAM-related preterm delivery.
Mol Hum Reprod 2001 Aug
PMID:NAD(P)H oxidase in human fetal membrane chorion laeve trophoblasts with or without chorioamnionitis: ultrastructural enzyme histochemical study. 1147 Aug 66


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