Dual oxidase 1, also known as DUOX1 or ThOX1 (for thyroid oxidase), is an enzyme which in humans is encoded by the DUOX1 gene. [5] DUOX1 was first identified in the mammalian thyroid gland. [6] In humans, two isoforms are found; hDUOX1 and hDUOX2. Human DUOX protein localization is not exclusive to thyroid tissue; hDUOX1 is prominent in airway epithelial cells and hDUOX2 in the salivary glands and gastrointestinal tract. [7] [8]
Investigations into reactive oxygen species (ROS) in biological systems have, until recently, focused on characterization of phagocytic cell processes. It is now well accepted that production of such species is not restricted to phagocytic cells and can occur in eukaryotic, non-phagocytic cell types via NADPH oxidase (NOX) or dual oxidase (DUOX). [9] [10] This new family of proteins, termed the NOX/DUOX family or NOX family of NADPH oxidases, consists of homologs to the catalytic moiety of phagocytic NADPH-oxidase, gp91phox. Members of the NOX/DUOX family have been found throughout eukaryotic species, including invertebrates, insects, nematodes, fungi, amoeba, alga, and plants (not found in prokaryotes). These enzymes clearly demonstrate regulated production of ROS as their sole function. Genetic analyses have implicated NOX/DUOX derived ROS in biological roles and pathological conditions including hypertension (NOX1), [11] innate immunity (NOX2/DUOX), [12] otoconia formation in the inner ear (NOX3), [13] and thyroid hormone biosynthesis (DUOX1/2). [14] The family currently has seven members including NOX1, NOX2 (formerly known as gp91phox), NOX3, NOX4, NOX5, DUOX1 (this enzyme) and DUOX2.
The current model for ROS generation by C. elegans DUOX1 (CeDUOX1) proposes that superoxide is generated through reduction of oxygen by two electrons extracted from oxidation of NADPH at the C-terminal NADPH oxidase domain. This unstable superoxide, generated at the extracellular surface, may rapidly convert to hydrogen peroxide and be utilized by the N-terminal peroxidase domain to facilitate tyrosine cross-linking. This model for CeDUOX1 activity was recently supported by a study of two point mutations localized within the peroxidase domain of CeDUOX1; G246D and D392N. [15] [16] Both mutations result in a blistering cuticle phenotype, resulting from the loss of tyrosine cross-linking activity. Neither mutant demonstrates a significant decrease in ROS production. These results suggest this peroxidase-like region is directly involved in enzymatic tyrosine cross-linking, but not responsible for ROS production.
Dual oxidases are characterized by a defining N-terminal, extracellular domain exhibiting considerable sequence identity with the mammalian peroxidases, a transmembrane (TM) segment appended to an EF-hand calcium-binding cytosolic region and a NOX2 homologous structure (six TMs tethered to NADPH oxidase). Topological studies place this peroxidase domain on the opposite side of the membrane from the NADPH oxidase domain.
hDUOX1 and hDUOX2 are 83% homologous, ~190 kDa in size (after extensive glycosylation contributing ~30 kDa in mass), and require maturation factors (DUOXA1 and DUOXA2) to achieve heterologous expression in full-length, active form. Mature DUOX enzymes produce H2O2; this activity is regulated by Ca2+ concentration through triggered dissociation of NOXA1 and possibly other as yet unidentified interacting proteins. [17] When sequence alignments were performed against other mammalian peroxidases, the histidine residues responsible for heme coordination were not conserved. [18] Due to this critical disparity, much speculation has surrounded the function of the DUOX peroxidase domain(s). Proposals for functionality include: superoxide dismutase activity, instead of peroxidase activity; a novel peroxidase mechanism; a protein-protein or Ca2+ induced conformational change which subsequently allows heme binding for peroxidase activity; or simply inactivity, as a vestigial domain.
Recent in vitro investigations into the ability of the DUOX1 domain to act as a peroxidase demonstrated that cell lysate from peroxidase expression in C. elegans and E. coli had tyrosine cross-linking activity. Further in vitro studies of human DUOX1 (hDUOX11-593) and C. elegans DUOX1 (CeDUOX11-589) were made possible by expression and purification via a baculovirus system. Evaluation of these proteins demonstrated that the isolated hDUOX11-593 does not bind heme and has no intrinsic peroxidase activity. In contrast, CeDUOX11-589 binds heme covalently and exhibits a modest peroxidase activity, but does not oxidize bromide ion. Surprisingly, the heme appears to have two covalent links to the C. elegans protein despite the absence of a second conserved carboxyl group in the active site. [19]
Two alternatively spliced transcript variants encoding the same protein have been described for this gene. [20]
Heme, or haem, is a ring-shaped iron-containing molecular component of hemoglobin, which is necessary to bind oxygen in the bloodstream. It is composed of four pyrrole rings with 2 vinyl and 2 propionic acid side chains. Heme is biosynthesized in both the bone marrow and the liver.
NADPH oxidase is a membrane-bound enzyme complex that faces the extracellular space. It can be found in the plasma membrane as well as in the membranes of phagosomes used by neutrophil white blood cells to engulf microorganisms. Human isoforms of the catalytic component of the complex include NOX1, NOX2, NOX3, NOX4, NOX5, DUOX1, and DUOX2.
NADPH oxidase 2 (Nox2), also known as cytochrome b(558) subunit beta or Cytochrome b-245 heavy chain, is a protein that in humans is encoded by the NOX2 gene. The protein is a superoxide generating enzyme which forms reactive oxygen species (ROS).
Nitric oxide dioxygenase (EC 1.14.12.17) is an enzyme that catalyzes the conversion of nitric oxide (NO) to nitrate (NO−
3) . The net reaction for the reaction catalyzed by nitric oxide dioxygenase is shown below:
Neutrophil cytosol factor 2 is a protein that in humans is encoded by the NCF2 gene.
Neutrophil cytosol factor 1, also known as p47phox, is a protein that in humans is encoded by the NCF1 gene.
NADPH oxidase 1 is an enzyme that in humans is encoded by the NOX1 gene.
NADPH oxidase 4 is an enzyme that in humans is encoded by the NOX4 gene, and is a member of the NOX family of NADPH oxidases.
Cytochrome b-245 light chain is a protein that in humans is encoded by the CYBA gene involved in superoxide production and phagocytosis.
Animal heme-dependent peroxidases is a family of peroxidases. Peroxidases are found in bacteria, fungi, plants and animals. On the basis of sequence similarity, a number of animal heme peroxidases can be categorized as members of a superfamily: myeloperoxidase (MPO); eosinophil peroxidase (EPO); lactoperoxidase (LPO); thyroid peroxidase (TPO); prostaglandin H synthase (PGHS); and peroxidasin.
Rac2 is a small signaling G protein, and is a member of the Rac subfamily of the family Rho family of GTPases. It is encoded by the gene RAC2.
Neutrophil cytosol factor 4 is a protein that in humans is encoded by the NCF4 gene.
Rho GDP-dissociation inhibitor 1 is a protein that in humans is encoded by the ARHGDIA gene.
Dual oxidase 2, also known as DUOX2 or ThOX2, is an enzyme that in humans is encoded by the DUOX2 gene. Dual oxidase is an enzyme that was first identified in the mammalian thyroid gland. In humans, two isoforms are found; hDUOX1 and hDUOX2. The protein location is not exclusive to thyroid tissue; hDUOX1 is prominent in airway epithelial cells and hDUOX2 in the salivary glands and gastrointestinal tract.
NADPH oxidase, EF-hand calcium binding domain 5, also known as NOX5, is a protein which in humans is encoded by the NOX5 gene.
NADPH oxidase organizer 1 is an enzyme that in humans is encoded by the NOXO1 gene.
NADPH oxidase 3 is an enzyme that in humans is encoded by the NOX3 gene.
NADPH oxidase activator 1 is an enzyme that in humans is encoded by the NOXA1 gene.
The respiratory tract antimicrobial defense system is a layered defense mechanism which relies on components of both the innate and adaptive immune systems to protect the lungs and the rest of the respiratory tract against inhaled microorganisms.
YedZ of E. coli has been examined topologically and has 6 transmembrane segments (TMSs) with both the N- and C-termini localized to the cytoplasm. von Rozycki et al. 2004 identified homologues of YedZ in bacteria and animals. YedZ homologues exhibit conserved histidyl residues in their transmembrane domains that may function in heme binding. Some of the homologues encoded in the genomes of various bacteria have YedZ domains fused to transport, electron transfer and biogenesis proteins. One of the animal homologues is the 6 TMS epithelial plasma membrane antigen of the prostate (STAMP1) that is over-expressed in prostate cancer. Some animal homologues have YedZ domains fused C-terminal to homologues of NADP oxidoreductases.
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