FRET-based detection of different conformations of MK2

MAP kinase-activated protein kinase 2 (MK2 or MAPKAP K2) is a stress-activated enzyme downstream to p38 MAPK. By fusion of green fluorescent protein variants 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 transition from an inactive/closed to an active/open conformation with an increase in the apparent distance between the fluorophores of similar to9 Angstrom. The closed conformation exists exclusively in the nucleus. Upon stress, the open conformation of MK2 rapidly becomes detectable in the cytoplasm and accumulates in the nucleus only when Crm1-dependent nuclear export is blocked. Hence, in living cells activation of MK2 and its nuclear export are coupled by a phosphorylation-dependent conformational switch.