| Assay Method Information | |
| | ERK5 Kinase Activity In Vitro Assay |
| Description: | Kinase activity was determined in an assay volume of 40 μl in kinase buffer (50 mM Tris-HCl, pH 7.5, 0.1 mM EGTA, 1 mM 2-mercaptoethanol) containing 200 ng of pure active ERK5 and the indicated amount of inhibitor. Reaction started by adding 10 mM magnesium acetate, and 50 μM [γ-32P]-ATP (500 cpm/pmol) and 250 μM PIMtide (ARKKRRHPSGPPTA) as substrates. Assays were carried out for 20 mM at 30° C., terminated by applying the reaction mixture onto p81 paper and the incorporated radioactivity measured as described previously. |
| Affinity data for this assay | |
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