Keywords

PKC, ABC Kinases, Phosphorylation, Thr, Ordered Phosphorylation, Protein Activation, Kinase Core Regulation

Reference

DOI: 10.1042/BJ20021626

Abstract

ABC kinases (Protein Kinases A, B/Akt, and C) share conserved phosphorylation motifs — activation loop, turn motif, and hydrophobic motif — that govern their maturation and signaling.
PKC serves as a paradigm illustrating how ordered phosphorylation events at these motifs regulate kinase maturation, activation, and down-regulation.
This review dissects how PDK-1-dependent and autophosphorylation events cooperate to control kinase structure and function, ensuring dynamic and reversible signaling.

Notes

1. ABC Kinases Share Conserved Phosphorylation Sites

  • Activation loop: Thr phosphorylation is essential for catalytic activation.
  • Turn motif: Anchors the C-terminal tail to stabilize kinase core, critical for maturation and structural stability.
  • Hydrophobic motif:
    • May dock onto the back of the kinase core, influencing C-helix positioning.
    • Docking site for PDK-1, facilitating activation loop phosphorylation.
  • Fancy insight: These three sites represent an “ordered phosphorylation code” regulating kinase activation and lifecycle.

2. The Life Cycle of PKC: Ordered Phosphorylation and Maturation

Step 1: Membrane Binding and Conformational Change

  • C1 and C2 domains bind membrane lipids (DAG, phospholipids), releasing pseudosubstrate from active site.
  • Activation loop becomes exposed, making PKC primed for phosphorylation.

Step 2: PDK-1 Phosphorylation

  • PDK-1 binds unphosphorylated hydrophobic motif, positioning itself to phosphorylate the activation loop.
  • Crucial step: Activation loop phosphorylation is rate-limiting and enables catalytic competence.

Step 3: Turn and Hydrophobic Motif Phosphorylation

  • After activation loop phosphorylation:
    • Turn motif phosphorylation stabilizes kinase core (possible autoP or by other kinases).
    • Hydrophobic motif phosphorylation further stabilizes the C-terminal tail, locks kinase in mature form.

Step 4: Mature, Catalytically Competent PKC

  • Thermally stable, protease- and phosphatase-resistant.
  • However, pseudosubstrate re-engagement may auto-inhibit PKC in the cytosol (inactive state).

3. Dephosphorylation, Hsp70 Stabilization, and PKC Recycling

  • Membrane-bound PKC: Active but susceptible to dephosphorylation, leading to degradation.
  • Turn motif dephosphorylation creates Hsp70 binding site, stabilizing deP-PKC.
  • Hsp70 allows refolding and re-phosphorylation, recycling PKC back into the functional pool.

Smart mechanism: PKC life cycle is maintained via balance of P, deP, and chaperone-mediated rescue.

4. PKC: A Model of Ordered Activation via Phosphorylation

  • Activation loop phosphorylation — prerequisite for activity.

  • Turn motif phosphorylation — structural stabilization, chaperone recruitment.

  • Hydrophobic motif phosphorylation — interface stabilization, regulation of kinase conformation.

  • Elegant hierarchy: Each phosphorylation event serves distinct yet coordinated roles in PKC activation and lifecycle.

5. Structural Coordination and Allosteric Regulation

  • C-terminal tail (C-tail) regulates kinase via three modules:
    • N-lobe tether (NLT): Positions C-helix, regulates active site access.
    • Active-site tether (AST): Interacts directly with ATP binding pocket.
    • C-lobe tether (CLT): Connects interlobe linker and αC–β4 loop — hinge for C-helix movement.
  • C-tail thus functions as cis-regulatory handle, modulated by trans-acting components.
  • Beautiful integration: C-tail and phosphorylation coordinate to regulate AGC kinases like PKC.

6. Conceptual Advances and Outstanding Questions

  • Ordered phosphorylation as a universal principle among AGC kinases (PKA, PKB, PKC).
  • PDK-1 as an essential priming kinase, positioning substrates correctly for activation.
  • Hydrophobic motif phosphorylation might order C-helix — a structural link to function.
  • Membrane interaction provides energy for opening autoinhibition — “binding energy drives activation”.
  • Still open:
    • How do other kinases like CaMK, CDPK regulate activation loop?
    • Is this membrane-mediated conformational switch a universal feature of all AGC-like kinases?

Take-home Messages

  • PKC activation is a multi-step phosphorylation-driven process, requiring careful coordination of activation loop, turn motif, and hydrophobic motif.
  • PDK-1 plays a central role in activation by phosphorylating the activation loop, but is also regulated via docking interactions.
  • The C-tail is a cis-acting module, modulating AGC kinase activity through structural interactions.
  • Dephosphorylation and chaperone stabilization (Hsp70) regulate PKC turnover and maintain kinase pools for rapid reuse.
  • PKC life cycle is a paradigm for ordered phosphorylation-dependent regulation — a blueprint for AGC kinases.

Final Thought: Activation through order, regulation through cycles — PKC and ABC kinases exemplify dynamic precision in cellular signaling.