Identification of a major determinant for serine-threonine kinase phosphoacceptor specificity.
Publication Type:Journal Article
Source:Mol Cell, Volume 53, Issue 1, p.140-7 (2014)
Keywords:Binding Sites, Crystallography, X-Ray, HEK293 Cells, Humans, Kinetics, p21-Activated Kinases, Peptides, Phosphorylation, Protein-Serine-Threonine Kinases, Substrate Specificity
<p>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. </p>