DETECTION OF ERK ACTIVATION BY A NOVEL MONOCLONAL-ANTIBODY

The mitogen-activated protein kinase, ERR is activated by a dual phosp horylation on threonine acid tyrosine residues. Using a synthetic diph ospho peptide, we have generated a monoclonal antibody directed to the active ERK. The antibody specifically identified the active doubly ph osphorylated, but not the inactive mono- or non- phosphorylated forms of ERKs, A direct correlation was observed between ERK activity and th e intensity in Western blot of mitogen-activated protein kinases from several species. The antibody was proven suitable for immunofluorescen ce staining, revealing a transient reactivity with ERKs that were tran slocated to the nucleus upon stimulation, In conclusion, the antibody can serve as a useful tool in the study of ERK signaling in a nide var iety of organisms. (C) 1997 Federation of European Biochemical Societi es.