The hydrophobic phosphorylation motif of conventional protein kinase C is regulated by autophosphorylation

Curr Biol. 1999 Jul 15;9(14):728-37. doi: 10.1016/s0960-9822(99)80332-7.


Background: A growing number of kinases are now known to be controlled by two phosphorylation switches, one on a loop near the entrance to the active site and a second on the carboxyl terminus. For the protein kinase C (PKC) family of enzymes, phosphorylation at the activation loop is mediated by another kinase but the mechanism for carboxy-terminal phosphorylation is still unclear. The latter switch contains two phosphorylation sites - one on a 'turn' motif and the second on a conserved hydrophobic phosphorylation motif - that are found separately or together in a number of other kinases.

Results: Here, we investigated whether the carboxy-terminal phosphorylation sites of a conventional PKC are controlled by autophosphorylation or by another kinase. First, kinetic analyses revealed that a purified construct of the kinase domain of PKC betaII autophosphorylated on the Ser660 residue of the hydrophobic phosphorylation motif in an apparently concentration-independent manner. Second, kinase-inactive mutants of PKC did not incorporate phosphate at either of the carboxy-terminal sites, Thr641 or Ser660, when expressed in COS-7 cells. The inability to incorporate phosphate on the hydrophobic site was unrelated to the phosphorylation state of the other key phosphorylation sites: kinase-inactive mutants with negative charge at Thr641 and/or the activation-loop position were also not phosphorylated in vivo.

Conclusions: PKC betaII autophosphorylates at its conserved carboxy-terminal hydrophobic phosphorylation site by an apparently intramolecular mechanism. Expression studies with kinase-inactive mutants revealed that this mechanism is the only one responsible for phosphorylating this motif in vivo. Thus, conventional PKC autoregulates the carboxy-terminal phosphorylation switch following phosphorylation by another kinase at the activation loop switch.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • COS Cells
  • Kinetics
  • Models, Biological
  • Molecular Sequence Data
  • Phosphorylation
  • Protein Kinase C / chemistry
  • Protein Kinase C / metabolism*
  • Serine / chemistry
  • Threonine / chemistry
  • Time Factors


  • Threonine
  • Serine
  • Protein Kinase C