Protein kinase specificity: role of primary structure elements and docking domains

Abstract

Protein phosphorylation, catalyzed by CDPKs and SNF1-related protein kinases (SnRK1s), is an important mechanism controlling the activity of metabolic enzymes and is often a component involved in signal transduction pathways. A major determinant of protein kinase specificity is generally considered to be the amino acid sequence surrounding the phosphorylated Ser/Thr. For both CDPKs and SnRK1s, the minimal phosphorylation motif is generally considered: f -x-Basic-x(2)-S/T (where f is a hydrophobic residue). The residues surrounding Ser158 of spinach sucrose-phosphate synthase (responsible for light/dark modulation of activity) match this motif. Most of the deduced sequences of SPS from dicot species surrounding the Ser158 regulatory phosphorylation site contain a proline residue at P-4 (where P is the phosphorylated serine); spinach is the exception and contains an arginine at P-4. This difference is significant, because a proline at P-4 selectively inhibits phosphorylation of the peptide substrate by CDPKs relative to SnRK1s. Thus, SPS in most dicot species may be selectively phosphorylated by SnRK1s (and not CDPKs) and thus may independent of calcium signaling. Recent results also suggest that `docking domains? (binding sites on target proteins that are removed from the phosphorylation site) may help localize protein kinases to their targets. Binding of the protein kinase PKIII (a SnRK1) to a putative docking site on sucrose-phosphate synthase has been demonstrated to specifically occur and may explain the observation that calcium-independent protein kinase activity co-immunoprecipitates with sucrose-phosphate synthase from leaf extracts. Collectively, the results indicate that protein kinase specificity may involve positive and negative recognition elements as well as putative docking domains.