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Enzyme catalysing the formation of the gasotransmitter NO(nitric oxide)

Nitric oxide synthases (NOSs) are a family of enzymes catalyzing the production of nitric oxide (NO) from L arginine. NO is an important cellular signaling molecule. It helps modulate vascular tone, insulin secretion, airway tone, and peristalsis, and is involved in angiogenesis and neural development. It may function as a retrograde neurotransmitter. Nitric oxide is mediated in mammals by the calcium calmodulin controlled isoenzymes eNOS (endothelial NOS) and nNOS (neuronal NOS). The inducible isoform, iNOS, involved in immune response, binds calmodulin at physiologically relevant concentrations, and produces NO as an immune defense mechanism, as NO is a free radical with an unpaired electron. It is the proximate cause of septic shock and may function in autoimmune disease. NOS catalyzes the reaction: NOSs are unique in that they use five cofactors and are the only known enzyme that binds flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN), heme, tetrahydrobiopterin (BH4) and calmodulin.

Arginine derived NO synthesis has been identified in mammals, fish, birds, invertebrates, and bacteria. 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). iNOS and nNOS are soluble and found predominantly in the cytosol, while eNOS is membrane associated. Evidence has been found for NO signaling in plants, but plant genomes are devoid of homologs to the superfamily which generates NO in other kingdoms.

As opposed to the critical calcium dependent regulation of constitutive NOS enzymes (nNOS and eNOS), iNOS has been described as calcium insensitive, likely due to its tight non covalent interaction with calmodulin (CaM) and Ca2+. The gene coding for iNOS is located on Chromosome 17. Induction of the high output iNOS usually occurs in an oxidative environment, and thus high levels of NO have the opportunity to react with superoxide leading to peroxynitrite formation and cell toxicity. These properties may define the roles of iNOS in host immunity, enabling its participation in anti microbial and anti tumor activities as part of the oxidative burst of macrophages. It has been suggested that pathologic generation of nitric oxide through increased iNOS production may decrease tubal ciliary beats and smooth muscle contractions and thus affect embryo transport, which may consequently result in ectopic pregnancy.

Endothelial NOS (eNOS), also known as nitric oxide synthase 3 (NOS3), generates NO in blood vessels and is involved with regulating vascular function. The gene coding for eNOS is located on Chromosome 7. eNOS localization to endothelial membranes is mediated by cotranslational N terminal myristoylation and post translational palmitoylation.

Bacterial NOS (bNOS) has been shown to protect bacteria against oxidative stress, diverse antibiotics, and host immune response. bNOS plays a key role in the transcription of superoxide dismutase (SodA). Bacteria late in the log phase who do not possess bNOS fail to upregulate SodA, which disables the defenses against harmful oxidative stress. Initially, bNOS may have been present to prepare the cell for stressful conditions but now seems to help shield the bacteria against conventional antimicrobials. As a clinical application, a bNOS inhibitor could be produced to decrease the load of Gram positive bacteria.

Nitric oxide synthases produce 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 Nω hydroxy L arginine (NOHLA) as an intermediate. 2 mol of O2 and 1.5 mol of NADPH are consumed per mole of NO formed. 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 reductase. They also share an amino terminal oxygenase domain containing a heme prosthetic group, which is linked in the middle of the protein to a calmodulin binding domain. Binding of calmodulin 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 oxygenase domain of each NOS isoform also contains an BH4 prosthetic group, which is required for the efficient generation of NO. Unlike other enzymes where BH4 is used as a source of reducing equivalents and is recycled by dihydrobiopterin reductase , BH4 activates heme bound O2 by donating a single electron, which is then recaptured to enable nitric oxide release. The first nitric oxide synthase to be identified was found in neuronal tissue (NOS1 or nNOS); the endothelial NOS (eNOS or NOS3) was the third to be identified. They were originally classified as "constitutively expressed" and "Ca2+ sensitive" but it is now known that they are present in many different cell types and that expression is regulated under specific physiological conditions. In NOS1 and NOS3, physiological concentrations of Ca2+ in cells regulate the binding of calmodulin to the "latch domains", thereby initiating electron transfer from the flavins to the heme moieties. In contrast, calmodulin remains tightly bound to the inducible and Ca2+ insensitive isoform (iNOS or NOS2) even at a low intracellular Ca2+ activity, acting essentially as a subunit of this isoform. Nitric oxide may itself regulate NOS expression and activity. Specifically, NO has been shown to play an important negative feedback regulatory role on NOS3, and therefore vascular endothelial cell function. This process, known formally as S nitrosation (and referred to by many in the field as S nitrosylation), has been shown to reversibly inhibit NOS3 activity in vascular endothelial cells. This process may be important because it is regulated by cellular redox conditions and may thereby provide a mechanism for the association between "oxidative stress" and endothelial dysfunction. In addition to NOS3, both NOS1 and NOS2 have been found to be S nitrosated, but the evidence for dynamic regulation of those NOS isoforms by this process is less complete. In addition, 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 may be particularly important after the induction of NOS2.

Ronopterin (VAS 203), also known as 4 amino tetrahydrobiopterin (4 ABH4), an analogue of BH4 (a cofactor of NOS), is an NOS inhibitor that is under development as a neuroprotective agent for the treatment of traumatic brain injury. Other NOS inhibitors that have been or are being researched for possible clinical use include cindunistat, A 84643, ONO 1714, L NOARG, NCX 456, VAS 2381, GW 273629, NXN 462, CKD 712, KD 7040, and guanidinoethyldisulfide, among others.