Keywords

CDPK, Calmodulin-like domain (CLD), Junction domain (JD), Calcium binding, EF-hand, NMR, Structural regulation

Reference

DOI: 10.1074/jbc.M311520200

Abstract

CDPKs are multi-domain, calcium-regulated kinases unique to plants and protozoa. Here, the high-resolution NMR structure of the CLD from soybean CDPK-alpha (with JD peptide) is presented. While the two lobes of CLD resemble EF-hand Ca2+-binding proteins, JD binding induces significant structural rearrangements, especially in the C-terminal lobe. Unexpectedly, CLD adopts an ‘open’ conformation, resembling calcineurin B rather than calmodulin, challenging functional assumptions based on sequence similarity.

Notes

1. Key Structural Insights

  • Both N- and C-terminal lobes form EF-hand helix-loop-helix structures.
  • C-terminal lobe interacts extensively with JD and is more defined structurally.
  • NMR mapping shows JD induces local structural changes, particularly in C-lobe.
  • Ca2+ stabilizes the NMR structure, but JD binding does not result in a stable complex — rather dynamic interaction.

The C-terminal lobe plays a central role in JD interaction, contrary to previous focus on N-lobe.

2. Structural Flexibility and Dynamic Features

  • Tether region (N-terminal of JD) is flexible, not rigid — important for regulating interaction dynamics.
  • Helix angles (EF-hand): slightly more closed, ~102–116°, suggesting tighter binding conformation.
  • Hydrophobic pockets identified on both lobes, surrounded by charged residues — essential for partner recognition and JD binding.

Structural flexibility may underlie CDPK’s unique regulation and specificity compared to calmodulin (CaM).

3. Binding and Functional Implications

  • JD binding induces open/closed state dynamics — two subpopulations observed:
    1. Side-by-side open state.
    2. Closed/compact state.
  • C-lobe remains flexible, adapting to JD presence.
  • A proposed “hinge” motion (around Tyr418 and Ala382) allows lobes to pivot and regulate access to binding partners.

Dynamic hinge-based mechanism — lobes are not independent like in CaM but coupled via pivot.

4. Comparative Structural Analysis

  • JD-CLD adopts a structure more similar to calcineurin B than CaM — open, not wrapping target.
  • Similarity with yeast and vertebrate CaM: transient lobe-lobe interactions.
  • Contrasts with CaM·CaM kinase complex, which tightly wraps the target.

CDPK evolved a distinct regulatory mode — hybrid of CaM and CNB features.

5. Mechanistic and Evolutionary Insights

  • CaM is constrained to interact with multiple partners, limiting evolutionary freedom.
  • CLD co-evolves with its own CDPK target, enabling high specificity and dynamic tuning.

CLD evolves as a unique regulator, perfectly adapted to its specific kinase — evolutionarily flexible.

  • Pivot/hinge-centered model:
    • C-lobe and N-lobe remain in contact, pivoting for regulation.
    • Diverges from CaM’s independent lobe model.

A new way to think about calcium-sensing modules: not independent, but tightly coupled and dynamically tuned.

Take-home Messages

  • CDPK’s CLD shows unexpected open conformation, similar to calcineurin B, not CaM.
  • C-lobe dominates interaction with JD, challenging traditional views of N-lobe dominance.
  • Dynamic hinge motion enables rapid conformational shifts upon Ca2+ binding and JD interaction.
  • CLD and JD interaction is highly specific, co-evolved for tailored regulation.
  • CDPK regulatory apparatus is an elegant, dynamic unit — a unique evolutionary solution to calcium signaling challenges in plants.

PDB References for Visualization

  • CLD structure: PDB: 1CDM, 1TCO — suggested for PyMOL visualization.