Purification notesab79586 was affinity purified from rabbit antiserum by affinity chromatography using epitope specific phosphopeptide. The antibody against non phosphopeptide was removed by chromatography using non phosphopeptide corresponding to the phosphorylation site.
FunctionDual specificity kinase. Is activated by cytokines and environmental stress in vivo. Catalyzes the concomitant phosphorylation of a threonine and a tyrosine residue in the MAP kinase p38.
Tissue specificityAbundant expression is seen in the skeletal muscle. It is also widely expressed in other tissues.
Involvement in diseaseNote=Defects in MAP2K3 may be involved in colon cancer.
Sequence similaritiesBelongs to the protein kinase superfamily. STE Ser/Thr protein kinase family. MAP kinase kinase subfamily. Contains 1 protein kinase domain.
Post-translational modificationsAutophosphorylated. Phosphorylation on Ser-218 and Thr-222 by MAP kinase kinase kinases regulates positively the kinase activity. Yersinia yopJ may acetylate Ser/Thr residues, preventing phosphorylation and activation, thus blocking the MAPK signaling pathway.
ICC/IF image of ab79586 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab79586, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, DyLight® 488 goat anti-rabbit IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Western blot - MEK3 (phospho T222) antibody (ab79586)
All lanes : Anti-MEK3 (phospho T222) antibody (ab79586) at 1/500 dilution
Lane 1 : Jurkat cell extracts treated
with serum (20 %, 15 minutes) Lane 2 : Jurkat cell extracts treated
with serum (20 %, 15 minutes) with immunizing peptide at 10 µg
Lysates/proteins at 30 µg per lane.
Predicted band size : 39 kDa Observed band size : 39 kDa Additional bands at : 32 kDa. We are unsure as to the identity of these extra bands.