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Mol Cell. 2014 Jan 9;53(1):140-7. doi: 10.1016/j.molcel.2013.11.013. Epub 2013 Dec 26.

Identification of a major determinant for serine-threonine kinase phosphoacceptor specificity.

Author information

1
Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA.
2
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520, USA; Department of Computer Science and Engineering, The Chinese University of Hong Kong, Shatin, New Territories, Hong Kong.
3
Division of Basic Science, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
4
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520, USA.
5
Donnelly Centre for Cellular and Biomolecular Research, University of Toronto, Toronto, ON M5S 3E1, Canada.
6
Oxford University, Nuffield Department of Clinical Medicine, Target Discovery Institute (TDI) and Structural Genomics Consortium (SGC), Oxford OX3 7FZ, UK; Ludwig Institute for Cancer Research, Old Road Campus Research Building, Oxford OX3 7DQ, UK.
7
Oxford University, Nuffield Department of Clinical Medicine, Target Discovery Institute (TDI) and Structural Genomics Consortium (SGC), Oxford OX3 7FZ, UK.
8
Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA. Electronic address: ben.turk@yale.edu.

Abstract

Eukaryotic protein kinases are generally classified as being either tyrosine or serine-threonine specific. Though not evident from inspection of their primary sequences, many serine-threonine kinases display a significant preference for serine or threonine as the phosphoacceptor residue. Here we show that a residue located in the kinase activation segment, which we term the "DFG+1" residue, acts as a major determinant for serine-threonine phosphorylation site specificity. Mutation of this residue was sufficient to switch the phosphorylation site preference for multiple kinases, including the serine-specific kinase PAK4 and the threonine-specific kinase MST4. Kinetic analysis of peptide substrate phosphorylation and crystal structures of PAK4-peptide complexes suggested that phosphoacceptor residue preference is not mediated by stronger binding of the favored substrate. Rather, favored kinase-phosphoacceptor combinations likely promote a conformation optimal for catalysis. Understanding the rules governing kinase phosphoacceptor preference allows kinases to be classified as serine or threonine specific based on their sequence.

PMID:
24374310
PMCID:
PMC3898841
DOI:
10.1016/j.molcel.2013.11.013
[Indexed for MEDLINE]
Free PMC Article

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