FRET-based detection of different conformations of MK2

MAP kinase-activated protein kinase 2 (MK2 or MAPKAP K2) is a stress-activa ted enzyme downstream to p38 MAPK. By fusion of green fluorescent protein v ariants to the N- and C-terminus we analysed conformational changes in the kinase molecule in vitro and in vivo. Activation of MK2 is accompanied by a decrease in fluorescence resonance energy transfer, indicating a transitio n from an inactive/closed to an active/open conformation with an increase i n the apparent distance between the fluorophores of similar to9 Angstrom. T he closed conformation exists exclusively in the nucleus. Upon stress, the open conformation of MK2 rapidly becomes detectable in the cytoplasm and ac cumulates in the nucleus only when Crm1-dependent nuclear export is blocked . Hence, in living cells activation of MK2 and its nuclear export are coupl ed by a phosphorylation-dependent conformational switch.