Summary: Nitric oxide synthase, oxygenase domain
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Nitric oxide synthase Edit Wikipedia article
The Nitric Oxide Synthases (NOS) are a group of enzymes responsible for the synthesis of [Nitric Oxide] from the terminal guanidino nitrogen atom of the semi-essential exclusive precursor amino acid L-Arginine in the presence of O2 and the cofactors Nicotinamide adenine dinucleotide phosphate (NADPH) and Flavin adenine dinucleotide (FAD) and tetrahydrobiopterin(BH4) and calmodulin. L-citrulline is a byproduct.
The different forms of NO synthase have been classified as
a. Neuronal NOS (nNOS or NOS1) which produces NO in neuronal tissue in both the central and peripheral nervous system. Neuronal NOS also performs a role in cell communication.
b. Inducible NOS (iNOS or NOS2) which can be found in the immune system but is also found in the cardiovascular system. It uses the oxidative stress of NO (a free radical) to be used by macrophages in immune defence against pathogens.
c. Endothelial NOS (eNOS or NOS3) generates NO in blood vessels and is involved with regulating vascular function. A constitutive Ca dependent NOS provides a basal release of NO.
All three isoforms (each of which is presumed to function as a homodimer during activation) share a carboxyl-terminal reductase domain homologous to the cytochrome p450 reductases. They also share an amino-terminal oxygenase domain containing a heme prosthetic group, which are linked in the middle of the protein by a calmodullin-binding domain. Binding of calmodullin appears to act as a "molecular switch" to enable electron flow from flavin prosthetic groups in the reductase domain to heme. This facilitates the conversion of O2 and L-arginine to NO and L-citrulline. The reductase domain of each NOS isoform also contains an H4B prosthetic group, which is required for the efficient generation of NO. Unlike other enzymes where H4B is used as a source of reducting equivalents and is recycled by dihyrobiopterin reductase H4B, H4B appears to be necessary to maintain a stable conformation for electron transport possible by promoting homodimerization.
The originally identified nitric oxide synthase was the NOS isoform identified in neuronal tissue known as nNOS or NOS1 followed by the endothelial NOS called eNOS or NOS3. They were originally classified as "constituitvely expressed" and "Ca2+ sensitive" but it is now known that they are present in may different cell types and that expression is regulated under specific physiological conditions.
In NOS1 (neuronal) and NOS3 (endothelial), physiological concentrations of Ca2+ in cells regulate the binding of calmodullin to the "latch domains" thereby initiating electron transfer from the flavins to the heme moieties. In contrast, calmodullin remains tightly bound to the inducible and Ca2+ insensitive isoform termed iNOS or NOS2 even at a low intracellular Ca2+ activity, acting essentially as a subunit of this isoform.
It is interesting that NO may itself regulate NOS expression and activity and has been shown to play an important negative feedback regulatory role on endothelial NO synthase, and therefore vascular endothelial cell function. Both NOS1 and NOS2 have been shown to form ferrous-nitrosyl complexes in their heme prosthetic groups that may act partially to self inactivate these enzymes under certain conditions. The rate-limiting step for the production of Nitric Oxide may well be the availability of L-arginine in some cell types. This is may particularly be important after the induction of NOS2
Nitric Oxide Release and Deactivation
Nitric Oxide generally exists as a lipophilic inorganic gas and is usually able to diffuse from producer to target cell. On reaching the vascular smooth muscle cells nitric oxide activates the soluble cyclic guanylate cyclase, which results in the formation of soluble cGMP. Nitric Oxide has an extremely short half-life, which has been estimated to be less than 4 seconds in biological solutions due to the rapid reaction with oxygen-derived free radicals, in particular the superoxide anion and oxyhaemoglobin. Nitric Oxide is rapidly oxidised by oxygenated haemoglobin to nitrite and then nitrate before being excreted in the urine. In addition to the direct effects of nitric oxide there is also evidence that it may well exert its effects through the formation of S-nitroso-thiols and metal-nitrosyl complexes, which can act as circulating reservoirs of nitric oxide.
The increase in soluble cGMP is matched with a decrease in intracellular calcium, which results in relaxation of the vascular smooth muscle. Nitric oxide can diffuse across the endothelial cell membrane to enter the adjacent vascular smooth muscle cells or alternatively pass into the lumen where it prevents platelet adhesion and aggregation by raising the level of cGMP in platelets. Nitric oxide also interacts with enzymes of the respiratory chain including aconitase and complex I and II and by this way can later tissue mitochondrial respiration.
Under resting conditions, it has been shown by studies on forearm blood flow that there is a continuous basal release of nitric oxide from the vascular endothelium. Infusing the NOS inhibitor L-NMMA into forearm vessels demonstrated a 50% fall in basal blood flow and attenuated the dilator response to infused acetylcholine but did not attenuate the vasodilatation due to glyceryl trinitrate. Basal release of NO is primarily due to shear stress or "viscous drag" which is determined by the vasoconstriction and flow rate and viscosity of the blood. This acts to balance the neurogenic and myogenic mediated vasoconstriction. The release of NO can be increased rapidly following the activation of specific receptors on endothelial cells resulting in an increased intracellular concentration of free calcium.
The other important physiological stimulants of NO release are factors released from aggregating platelets, such as serotonin and adenosine nucleotides. This strong release of NO upon exposure to platelet products and thrombin is of crucial importance in preventing unwarranted intravascular coagulation assuming the endothelium is intact.
The most important role of nitric oxide as we have discussed is in the control of vascular tone. Aside from the response to acetylcholine, the release of NO is stimulated by shear stress, serotonin and ADP as mentioned above. Other stimulants include thrombin and histamine. Only a few vasodilators work independently of the endothelium, most notably the nitrovasodilators such as nitroprusside and nitroglycerine, and other mediators such as prostacyclin and adenosine
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Nitric oxide synthase, oxygenase domain Provide feedback
No Pfam abstract.
Crane BR, Arvai AS, Ghosh DK, Wu C, Getzoff ED, Stuehr DJ, Tainer JA; , Science 1998;279:2121-2126.: Structure of nitric oxide synthase oxygenase dimer with pterin and substrate. PUBMED:9516116 EPMC:9516116
External database links
This tab holds annotation information from the InterPro database.
InterPro entry IPR004030
This entry represents the N-terminal of the nitric oxide synthases.
Nitric oxide synthase ( EC ) (NOS) enzymes produce nitric oxide (NO) by catalysing a five-electron oxidation of a guanidino nitrogen of L-arginine (L-Arg). Oxidation of L-Arg to L-citrulline occurs via two successive monooxygenation reactions producing N(omega)-hydroxy-L-arginine as an intermediate. 2 mol of O(2) and 1.5 mol of NADPH are consumed per mole of NO formed [ PUBMED:8782597 ].
Arginine-derived NO synthesis has been identified in mammals, fish, birds, invertebrates, plants, and bacteria [ PUBMED:8782597 ]. Best studied are mammals, where three distinct genes encode NOS isozymes: neuronal (nNOS or NOS-1), cytokine-inducible (iNOS or NOS-2) and endothelial (eNOS or NOS-3) [ PUBMED:7510950 ]. iNOS and nNOS are soluble and found predominantly in the cytosol, while eNOS is membrane associated. The enzymes exist as homodimers, each monomer consisting of two major domains: an N-terminal oxygenase domain, which belongs to the class of haem-thiolate proteins, and a C-terminal reductase domain, which is homologous to NADPH:P450 reductase ( EC ). The interdomain linker between the oxygenase and reductase domains contains a calmodulin (CaM)-binding sequence. NOSs are the only enzymes known to simultaneously require five bound cofactors animal NOS isozymes are catalytically self-sufficient. The electron flow in the NO synthase reaction is: NADPH --> FAD --> FMN --> haem --> O(2).
eNOS localisation to endothelial membranes is mediated by cotranslational N-terminal myristoylation and post-translational palmitoylation [ PUBMED:9199168 ]. The subcellular localisation of nNOS in skeletal muscle is mediated by anchoring of nNOS to dystrophin. nNOS contains an additional N-terminal domain, the PDZ domain [ PUBMED:7535955 ]. Some bacteria, like Bacillus halodurans, Bacillus subtilis or Deinococcus radiodurans, contain homologues of NOS oxygenase domain.
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|Molecular function||nitric-oxide synthase activity (GO:0004517)|
|Biological process||nitric oxide biosynthetic process (GO:0006809)|
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|Seed source:||Structural domain|
|Number in seed:||66|
|Number in full:||2340|
|Average length of the domain:||326.8 aa|
|Average identity of full alignment:||53 %|
|Average coverage of the sequence by the domain:||37.91 %|
|HMM build commands:||
build method: hmmbuild -o /dev/null HMM SEED
search method: hmmsearch -Z 61295632 -E 1000 --cpu 4 HMM pfamseq
|Family (HMM) version:||18|
|Download:||download the raw HMM for this family|
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For those sequences which have a structure in the Protein DataBank, we use the mapping between UniProt, PDB and Pfam coordinate systems from the PDBe group, to allow us to map Pfam domains onto UniProt sequences and three-dimensional protein structures. The table below shows the structures on which the NO_synthase domain has been found. There are 1326 instances of this domain found in the PDB. Note that there may be multiple copies of the domain in a single PDB structure, since many structures contain multiple copies of the same protein sequence.
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AlphaFold Structure Predictions
The list of proteins below match this family and have AlphaFold predicted structures. Click on the protein accession to view the predicted structure.