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Literature summary for 1.14.13.39 extracted from
- N-Glycosylation at Asn695 might suppress inducible nitric oxide synthase activity by disturbing electron transfer (2020), Acta Biochim. Biophys. Sin., 52, 1360-1372 .
Activating Compound
| Activating Compound | Comment | Organism | Structure |
|---|---|---|---|
| Calmodulin | required | Mus musculus |  |
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| gene nos2, recombinant expression of wild-type and mutant enzymes in HEK293 cells or Escherichia coli | Mus musculus |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| N695Q | site-directed mutagenesis, mutating Asn695 to Gln695 yields an iNOS that exhibits greater enzyme activity compared to wild-type. NO produced by N695Q iNOS-transformed HEK293 cells is 1.32fold greater than that of N-glycosylated iNOS, the increased enzyme activity of N695Q iNOS in HEK293 cells was caused by loss of N-glycan | Mus musculus |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| Nomega-nitro-L-arginine methylester | L-NAME, an inhibitor of iNOS | Mus musculus |
Localization
| Localization | Comment | Organism | GeneOntology No. | Textmining |
|---|---|---|---|---|
| cell membrane | - |
Mus musculus | - |
- |
| cytoplasm | - |
Mus musculus | 5737 | - |
| endoplasmic reticulum | iNOS contains several characteristic endoplasmic reticulum (ER)-localization sequences. iNOS is likely to be glycosylated in the endoplasmic reticulum | Mus musculus | 5783 | - |
| Golgi apparatus | - |
Mus musculus | 5794 | - |
| additional information | N-glycosylation of proteins starts in the endoplasmic reticulum, continues in the Golgi, and ends at the plasma membrane. As mentioned above, iNOS has characteristic ER localization sequences and locates in the Golgi and cell membrane. iNOS is located both in cytoplasm and on cell membrane, it is not only a cytoplasmic protein but also a membrane protein | Mus musculus | - |
- |
Metals/Ions
| Metals/Ions | Comment | Organism | Structure |
|---|---|---|---|
| Ca2+ | required | Mus musculus | |
| Fe2+ | in the heme | Mus musculus | |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| 2 L-arginine + 3 NADPH + 3 H+ + 4 O2 | Mus musculus | overall reaction | 2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O | - |
? | |
| 2 L-arginine + 3 NADPH + 3 H+ + 4 O2 | Mus musculus C57BL/6 | overall reaction | 2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O | - |
? | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Mus musculus | P29477 | - |
- |
| Mus musculus C57BL/6 | P29477 | - |
- |
Posttranslational Modification
| Posttranslational Modification | Comment | Organism |
|---|---|---|
| glycoprotein | iNOS is N-glycosylated in vitro and in vivo. Mass spectrometry studies identify Asn695 as an N-glycosylation site of murine iNOS, mapping of the iNOS N-glycosylation site. iNOS expressed by LPS- and IFN-gamma-stimulated Raw-264.7 cells can be recognized by concanavalin A (ConA). Glycosylation at Asn695 might suppress inducible nitric oxide synthase activity by disturbing electron transfer | Mus musculus |
Source Tissue
| Source Tissue | Comment | Organism | Textmining |
|---|---|---|---|
| RAW-264.7 cell | - |
Mus musculus | - |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| 2 L-arginine + 3 NADPH + 3 H+ + 4 O2 | overall reaction | Mus musculus | 2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O | - |
? | |
| 2 L-arginine + 3 NADPH + 3 H+ + 4 O2 | overall reaction | Mus musculus C57BL/6 | 2 L-citrulline + 2 nitric oxide + 3 NADP+ + 4 H2O | - |
? | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| dimer | all three NOS isoforms are composed of an N-terminal catalytic oxygenase module (NOSox) and a C-terminal-electron-supplying reductase module (NOSred), and NOSox and NOSred are linked by a Ca2+/calmodulin binding region. NOSox contains a heme-binding site, a tetrahydrobiopterin-binding site and an arginine-binding site. NOSred contains an flavin mononucleotide-binding site, an flavin adenine dinucleotide-binding site and an NADPH-binding site. Only dimerized NOS has catalytic activity | Mus musculus |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| inducible nitric oxide synthase | - |
Mus musculus |
| iNOS | - |
Mus musculus |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 37 | - |
assay at | Mus musculus |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7.4 | - |
assay at | Mus musculus |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| FAD | - |
Mus musculus | |
| FMN | - |
Mus musculus | |
| heme | - |
Mus musculus | |
| NADPH | - |
Mus musculus | |
General Information
| General Information | Comment | Organism |
|---|---|---|
| malfunction | attachment of N-glycan to the Asn695 residue inhibits activity by disturbing electron transfer. N-glycosylated iNOS consumes NADPH more slowly than the unliganded enzyme. Mutating Asn695 to Gln695 yields an iNOS that exhibits greater enzyme activity compared to wild-type. NO produced by N695Q iNOS-transformed HEK293 cells is 1.32fold greater than that of N-glycosylated iNOS, the increased enzyme activity of N695Q iNOS in HEK293 cells was caused by loss of N-glycan | Mus musculus |
| additional information | residue Asn695 of the mouse iNOS locates at the hinge segment which connects the FMN-binding domain to the FAD-binding domain. The electrostatic and flexibility properties of hinge segment are critical for electron transfer from CPR to its redox partners. For mouse iNOS, N-glycosylation of Asn695 might disturb electron transfer by influencing the electrostatic and flexibility properties of the hinge segment | Mus musculus |
| physiological function | inducible nitric oxide synthase (iNOS) is a key inflammatory factor. It functions in both acute and chronic inflammation. Nitric oxide (NO) is a signaling mediator with many diverse and often contradictory biological activities. In mammals, NO is produced by a family of nitric oxide synthase (NOS). The NOS family includes neuronal nitric oxide synthase (nNOS, type I), inducible nitric oxide synthase (iNOS, type II), and endothelial nitric oxide synthase (eNOS, type III). All these three NOS isoforms catalyze a similar reaction. Consuming NADPH and O2, NOS oxidizes L-arginine into L-citrulline and releases NO. The reaction is an oxidation-reduction reaction, and electron transfer plays a vital role. Inducible nitric oxide synthase (iNOS) plays critical roles in the inflammatory response and host defense. The essence of nitric oxide synthase catalytic reaction is an electron transfer process, which involves a series of conformational changes, and the linker between the flavin mononucleotide-binding domain and the flavin adenine dinucleotide-binding domain plays vital roles in the conformational changes. Residue Asn695 is part of the linker. Enzyme iNOS is N-glycosylated at its Asn695 residue and N-glycosylation of Asn695 might suppress iNOS activity by disturbing electron transfer | Mus musculus |
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