A PEF/Y substrate recognition and signature motif plays a critical role in DAPK-related kinase activity.
Knowledge about protein kinase substrate preferences is biased toward residues immediately adjacent to the site of phosphorylation. By a combined structural, biochemical, and cellular approach, we have discovered an unexpected substrate recognition element with the consensus sequence PEF/Y in the tumor suppressor death-associated protein kinase 1. This motif can be effectively blocked by a specific pseudosubstrate-type interaction with an autoregulatory domain of this kinase. In this arrangement, the central PEF/Y glutamate interacts with a conserved arginine distant to the phosphorylation site in sequence and structure. We also demonstrate that the element is crucial for kinase activity regulation and substrate recognition. The PEF/Y motif distinguishes close death-associated protein kinase relatives from canonical calcium/calmodulin-dependent protein kinases. Insight into this signature and mode of action offers new opportunities to identify specific small molecule inhibitors in PEF/Y-containing protein kinases.
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany; European Molecular Biology Laboratory Heidelberg, Meyerhofstrasse 1, 69117 Heidelberg, Germany.
Koen
K
Temmerman
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany.
Iñaki
I
de Diego
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany.
Vivian
V
Pogenberg
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany.
Bertrand
B
Simon
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany.
Weronika
W
Jonko
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany; European Molecular Biology Laboratory Heidelberg, Meyerhofstrasse 1, 69117 Heidelberg, Germany.
Xun
X
Li
European Molecular Biology Laboratory Hamburg, Notkestrasse 85, 22603 Hamburg, Germany. Electronic address: wilmanns@embl-hamburg.de.
Matthias
M
Wilmanns
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D018360:Crystallography, X-Ray; D064550:Death-Associated Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment; D013379:Substrate Specificity
D010455:Peptides; C579096:DAPK1 protein, human; D064550:Death-Associated Protein Kinases