An SH3 domain‐mediated interaction between the phagocyte NADPH oxidase factors p40phox and p47phox

Ito, Takashi; Nakamura, Rika; Sumimoto, Hideki; Takeshige, Koichiro; Sakaki, Yoshiyuki

doi:10.1016/0014-5793(96)00387-0

Cited by 51 publications

(53 citation statements)

References 22 publications

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“…Domain pr, revealed by the binding assay, corresponds closely to the C-terminal proline-rich motif, shown to interact with the C-terminal SH3 domain in p67 phox (31,35,47,67,68). This tail-to-tail interaction is not involved in NADPH oxidase activation in the cell-free system.…”

Section: P47mentioning

confidence: 88%

Mapping of Functional Domains in p47 Involved in the Activation of NADPH Oxidase by “Peptide Walking”

Morozov

Lotan

Joseph

et al. 1998

Journal of Biological Chemistry

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The superoxide generating NADPH oxidase of phagocytes consists, in resting cells, of a membrane-associated electron transporting flavocytochrome (cytochrome b 559 ) and four cytosolic proteins as follows: p47 phox , p67 phox , p40 phox , and the small GTPase, Rac(1 or 2). Activation of the oxidase is consequent to the assembly of a membrane-localized multimolecular complex consisting of cytochrome b 559 and the cytosolic components. We used "peptide walking" (Joseph, G., and Pick, E. (1995) J. Biol. Chem. 270, 29079 -29082) for mapping domains in the amino acid sequence of p47 phox participating in the molecular events leading to the activation of NADPH oxidase. Ninety-five overlapping pentadecapeptides, with a four-residue offset between neighboring peptides, spanning the complete p47 phox sequence, were tested for the ability to inhibit NADPH oxidase activation in a cell-free system. This consisted of solubilized macrophage membranes, recombinant p47 phox , p67 phox , and Rac1, and lithium dodecyl sulfate, as the activator. Eight functional domains were identified and labeled a-h. phox , and domains f and h, in the C-terminal half, represent newly identified entities, for which there is no earlier experimental evidence of involvement in NADPH oxidase activation. "Peptide walking" was also applied to the identification of domains in p47 phox mediating binding to p67 phox . This was done by quantifying, by enzyme-linked immunosorbent assay, the binding of p67 phox , in solution, to a series of 95 overlapping biotinylated p47 phox peptides, attached to streptavidin-coated 96-well plates. A single proline-rich domain (residues 357-371) was found to bind p67 phox in the absence and presence of lithium dodecyl sulfate.Phagocytic cells produce, in response to appropriate stimuli, a variety of oxygen-derived toxic radicals, all of which are derived from superoxide (O 2 . ). 1 O 2 . is generated by the NADPHderived one-electron reduction of molecular oxygen, catalyzed by a membrane-associated heterodimeric flavocytochrome (cytochrome b 559 ), composed of a 91-kDa glycoprotein (gp91 phox ) and a 22-kDa protein (p22 phox ), and incorporating two redox centers, FAD and two hemes (reviewed in Refs. 1-3). The conversion of cytochrome b 559 from the resting to the activated state is, most likely, the result of a conformational change leading to a high turnover electron flow from NADPH to oxygen, through the redox centers. This change in conformation is brought about by protein-protein interactions involving the two cytochrome b 559 subunits and four cytosolic regulatory proteins, p47phox , p67 phox , p40 phox , and the small GTPase Rac(1 or 2) (reviewed in Refs. 4 and 5). It is commonly assumed that activation leads to the formation of a membrane-localized multimolecular structure, known as the NADPH oxidase complex. The physical expression of this, in the intact cell, is the translocation of parts of p47 phox and p67 phox to the plasma membrane (6) associated with the phosphorylation of p47 phox at multiple sites (7). Whe...

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Section: P47mentioning

confidence: 88%

Mapping of Functional Domains in p47 Involved in the Activation of NADPH Oxidase by “Peptide Walking”

Morozov

Lotan

Joseph

et al. 1998

Journal of Biological Chemistry

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“…2). The SH3 domain of p40 phox can compete with the C-terminal SH3 domain of p67 phox for binding to the proline rich region of p47 phox [66,67] [79][80][81][82][83][84][85]. A large number of isologs have now been catalogued in species ranging from plants, fungi, amoeba, nematodes, insects, and vertebrates.…”

Section: Domain Structures Of the Phox Regulatory Subunitsmentioning

confidence: 99%

Regulation of Nox and Duox enzymatic activity and expression

Lambeth

Kawahara

Diebold

2007

Free Radical Biology and Medicine

449

418

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SummaryIn recent years, it has become clear that reactive oxygen species (ROS, which include superoxide, hydrogen peroxide and other metabolites) are produced in biological systems. Rather than being simply a byproduct of aerobic metabolism, it is now recognized that specific enzymes ---the Nox (NADPH-oxidase) and Duox (Dual oxidase) enzymes ----seem to have the sole function of generating ROS in a carefully regulated manner, and key roles in signal transduction, immune function, hormone biosynthesis and other normal biological functions are being uncovered. The prototypical Nox is the respiratory burst oxidase or phagocyte oxidase, which generates large amounts of superoxide and other reactive species in the phagosomes of neutrophils and macrophages, playing a central role in innate immunity by killing microbes. This enzyme system has been extensively studied over the past two decades, and provides a basis for comparison with the more recently described Nox and Duox enzymes, which generate ROS in a variety of cells and tissues. This review first considers the structure and regulation of the respiratory burst oxidase, and then reviews recent studies relating to the regulation of the activity of the novel Nox/Duox enzymes. The regulation of Nox and Duox expression in tissues and by specific stimuli is also considered here. An accompanying review considers biological and pathological roles of the Nox family of enzymes. The Respiratory Burst Oxidase of Phagocytes a. The respiratory burstThe "respiratory burst" refers to the early observation that when professional phagocytes such as neutrophils and macrophages are exposed to microbes, they consume large amounts of oxygen. Unexpectedly, this oxygen consumption was not inhibited by cyanide, an inhibitor of mitochondrial electron transport. This observation led to a more than 25 year search for the enzymatic origin of the respiratory burst and to the eventual discovery and molecular characterization of the phagocytic NADPH-oxidase or "respiratory burst oxidase". The phagocyte oxidase generates superoxide via the one electron-reduction of oxygen by NADPH, with secondary production of hydrogen peroxide, HOCl and other activated forms of oxygen. Together, these reactive oxygen species (ROS) participate in host defense by killing invading microbes. b. gp91 phox , the catalytic moiety of the respiratory burst oxidaseThe phagocytic NADPH-oxidase consists of a membrane-localized glycosylated, catalytic subunit, gp91 phox (which has also come to be known as Nox2, a terminology that will be used in this review), along with a second membrane-associated subunit, p22 phox , both depicted in Fig. 1. Nox2 and p22 phox stabilize one another in a tightly associated heterodimer which is referred to as flavocytochrome b 558 . The C-terminal half of Nox2 forms a domain that is Contact information: 148 Whitehead Biomedical Research Building, Department of Pathology and Laboratory Medicine, 615 Michael Street, Atlanta, GA 30322, U.S. A. Phone: 404-727-5875, Fax: 404-712-2979, e-mail: nox...

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“…phox C terminus (17)(18)(19), and p67 phox PB1 domain (20). The physiological role of p40 phox is still a matter of debate.…”

mentioning

confidence: 99%

Effects of p47 C Terminus Phosphorylations on Binding Interactions with p40 and p67

Massenet

Chenavas

Cohen‐Addad

et al. 2005

Journal of Biological Chemistry

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The neutrophil NADPH oxidase produces superoxide anions in response to infection. This reaction is activated by association of cytosolic factors, p47 phox and p67 . These data reveal that SH3 p40 can interact with a consensus polyproline motif but also with a noncanonical motif of the p47 phox C terminus. The electrostatic surfaces of both SH3 are very different, and therefore the binding preference for C-SH3 p67 can be attributed to the polyproline motif recognition and particularly to the Arg-368 p47 binding mode. The noncanonical motif contributes equally to interaction with both SH3. The influence of serine phosphorylation on residues 359/ 370 and 379 on the affinity for both SH3 domains has been checked. We conclude that contrarily to previous suggestions, phosphorylation of Ser-359/370 does not modify the SH3 binding affinity for both SH3, whereas phosphorylation of Ser-379 has a destabilizing effect on both interactions. Other mechanisms than a phosphorylation induced switch between the two SH3 must therefore take place for NADPH oxidase activation cascade to start.

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An SH3 domain‐mediated interaction between the phagocyte NADPH oxidase factors p40^phox and p47^phox

Cited by 51 publications

References 22 publications

Mapping of Functional Domains in p47 Involved in the Activation of NADPH Oxidase by “Peptide Walking”

Mapping of Functional Domains in p47 Involved in the Activation of NADPH Oxidase by “Peptide Walking”

Regulation of Nox and Duox enzymatic activity and expression

Effects of p47 C Terminus Phosphorylations on Binding Interactions with p40 and p67

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