Catalog # NP4091

iNOS (Tyr-1055)[conserved site], phospho-specific Antibody

Rabbit Polyclonal

Application / Dilution

ELISA	1:2000
ICC	1:400
WB	1:1000

Size 100 μl

Species Reactivity Hu, Rt, Ms

MW 130 kDa

Nitric oxide synthases (NOS), the enzymes responsible for synthesis of NO, are homodimers whose monomers are themselves two fused enzymes: a cytochrome reductase and a cytochrome that requires three cosubstrates (L-arginine, NADPH, and oxygen) and five cofactors or prosthetic groups (FAD, FMN, calmodulin, tetrahydrobiopterin, and heme). Several distinct NOS isoforms are produced from three distinct genes, inducible NOS (iNOS, NOS-II), neuronal NOS (bNOS, NOS-I), and endothelial NOS (eNOS, ecNOS, NOS-III). The inducible form of NOS, iNOS, is Ca2+ independent and is expressed in a broad range of cell types in response to stimulation with cytokines and exposure to microbial products. Phosphorylation of iNOS may regulate its activity and stability. Src kinase-induced phosphorylation of Tyr-1055 in iNOS reduces proteasomal degradation of iNOS, leading to increased NO production. Thus, phosphorylation may be an additional control for regulating iNOS activity in response to inflammatory conditions.

References

Tyryshkin, A. et al. (2009) J Biol Chem. 285(1):784.
Kleinert, H. et al. (2003) Biol Chem. 384(10-11):1343.
Xie, Q.W. et al. (1992) Science 256:225.

Western blot analysis of mouse macrophages (J774A.1) treated with LPS (1µg/ml) for 18 hrs followed by pervanadate (1 mM) for 30 min. (lanes 1, 3 & 5). The blots were then treated with akaline phosphatase (lanes 2, 4 & 6). Blots were probed with rabbit polyclonal anti-inducible Nitric Oxide Synthase (iNOS) (lanes 1 & 2), anti-iNOS (Tyr-1055) (lanes 3 & 4), and mouse monoclonal anti-iNOS (lanes 5 & 6).

Immunocytochemical labeling of nNOS phosphorylation in rat PC12 cells differentiated with NGF. 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 appropriate secondary antibody conjugated to DyLight® 594.

Phospho-iNOS (Tyr-1055) synthetic peptide (coupled to carrier protein) corresponds to amino acids surrounding Tyr-1055 in human iNOS. This sequence is well conserved in rat and mouse iNOS, and has some homology to the conserved sites in eNOS (Tyr-1087) and nNOS (Tyr-1326).

*For more information, see UniProt Accession P35228

Rabbit polyclonal, affinity-purified antibody is supplied in 100µl phosphate-buffered saline, 50% glycerol, 1 mg/ml BSA, and 0.05% sodium azide. Store at –20°C. Stable for 1 year.

The products are are safely shipped at ambient temperature for both domestic and international shipments. Each product is guaranteed to match the specifications as indicated on the corresponding technical data sheet. Please store at -20C upon arrival for long term storage.

The antibody detects a 130 kDa* protein corresponding to induced NOS on SDS-PAGE immunoblots of mouse macrophages treated with LPS followed by pervanadate. The antibody also detects a 150 kDa doublet that corresponds with higher molecular weight forms of NOS, which are also observed using other anti-iNOS antibodies. In addition, the antibody detects a non-specific 120 kDa band that has not been identified.

*All molecular weights (MW) are confirmed by comparison to Bio-Rad Rainbow Markers and to western blot mobilities of known proteins with similar MW.

This kit contains: