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FRET‐based detection of different conformations of MK2
Author(s) -
Neininger Armin,
Thielemann Heiko,
Gaestel Matthias
Publication year - 2001
Publication title -
embo reports
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 4.584
H-Index - 184
eISSN - 1469-3178
pISSN - 1469-221X
DOI - 10.1093/embo-reports/kve157
Subject(s) - förster resonance energy transfer , nucleus , cytoplasm , chemistry , biophysics , phosphorylation , nuclear export signal , protein kinase a , fusion protein , microbiology and biotechnology , kinase , conformational change , in vitro , green fluorescent protein , p38 mitogen activated protein kinases , enzyme , biochemistry , fluorescence , cell nucleus , biology , recombinant dna , gene , physics , quantum mechanics
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 ∼9 Å. 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.