Nitric oxide (NO) has a broad range of biological activities and is implicated in many cell signaling pathways. Nitric oxide synthases (NOS), the enzymes responsible for synthesis of NO, are fused enzymes that include a cytochrome reductase and a cytochrome that requires three cosubstrates (L-arginine, NADPH, and oxygen) and five cofactors (FAD, FMN, calmodulin, tetrahydrobiopterin, and heme). The NOS family includes two constitutive Ca2+/CaM-dependent forms of NOS: nNOS (also designated bNOS, NOS-I), whose activity was first identified in neurons and eNOS (also designated ecNOS, NOS-III) first identified in endothelial cells. The inducible form of NOS, iNOS (also designated NOS-II), is Ca2+ independent and is induced by microbial products in various cell types. Phosphorylation of NOS enzymes may regualte their activity. For example, Src kinase-induced phosphorylation of Tyr-1055 in iNOS reduces proteasomal degradation of iNOS, while shear stress induced phosphorylation of Tyr-657 in eNOS attenuates enzyme activity. Both of these tyrosine sites are conserved in nNOS, and may have important roles for regulate neuronal NOS activity.
References
Fisslthaler, B. et al. (2008) Circ Res. 102:1520.
Tyryshkin, A. et al. (2009) J Biol Chem. 285(1):784.
Loot, A. E. et al. (2009) J Exp Med. 206(13):2889.
Western blot analysis of nNOS expression in adult mouse brain (lanes 1 & 3) and rat GC cells (lanes 2 & 4). The blots were probed with mouse monoclonal anti-nNOS (C-terminal region) at 1:1000 (lanes 1 & 2) or rabbit polyclonal anti-nNOS at 1:250 (lanes 3 & 4).
Labeling of nNOS phosphorylation in rat PC12 cells. The cells were probed with mouse monoclonal (mAb) nNOS (NM4011), and rabbit polyclonal (pAb) nNOS (C-terminal region), nNOS (Tyr-895)/eNOS (Tyr-657), and nNOS (Tyr-1326)/iNOS (Tyr-1055). The antibodies were detected using secondary antibody conjugated to DyLight® 594.
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*All molecular weights (MW) are confirmed by comparison to Bio-Rad Rainbow Markers and to western blot mobilities of known proteins with similar MW.
Product References:
RS3251 Kawasaki, H. et al. (2013) World J Gastroenter. 19(17):2629. WB, ICC: mouse intestinal myofibroblastsNM4011 Stasko, S. et al. (2013) J Appl Physiol. 114(11):1629. WB: mouse diaphragm, normal, deficient
RS3251 Estrada-Bernal, A. et al. (2011) J Neurooncol. 102:353. Western Blot
MS3001 Estrada-Bernal, A. et al. (2011) J Neurooncol. 102:353. Western Blot
This kit contains:
| CATALOG# | DESCRIPTION | SIZE | APPLICATIONS | SPECIES REACTIVITY | MW (kDa) |
| eNOS (Tyr-657)/nNOS (Tyr-895), phospho-specific Rabbit pAb | 50 μl | WB, E, ICC | Hu, Rt, Ms | 140 | |
| iNOS (Tyr-1055)[conserved site], phospho-specific Rabbit pAb | 50 μl | WB, E, ICC | Hu, Rt, Ms, Ck | 130 | |
| nNOS (C-terminal region) Mouse mAb | 50 μl | WB, E, ICC, IHC | Hu, Rt, Ms | 155 | |
| nNOS (C-terminal regiom) Rabbit pAb | 50 μl | WB, E, ICC, IHC | Hu, Rt, Ms | 155 | |
| Anti-Mouse Ig:HRP Donkey pAb | 100 μl | WB, E | Ms | ||
| Anti-Rabbit Ig Light ChainSpecific:HRP Mouse mAb | 100 μl | WB, E, ICC, IHC | Rb |
KIT SUMMARY
The nNOS phospho-regulation kit can be used to examine phosphorylation of nNOS (Tyr-895)/eNOS (Tyr-657) and nNOS (Tyr-1326)/iNOS (Tyr-1055) conserved sites relative to the total expression of nNOS. The kit includes rabbit polyclonal and mouse monoclonal antibodies to detect total nNOS, and secondary reagents for rabbit polyclonal and mouse monoclonal antibody detection.
