The C-terminal random coil region tunes the Ca²⁺-binding affinity of S100A4 through conformational activation

PLoS One. 2014 May 15;9(5):e97654. doi: 10.1371/journal.pone.0097654. eCollection 2014.

Abstract

S100A4 interacts with many binding partners upon Ca2+ activation and is strongly associated with increased metastasis formation. In order to understand the role of the C-terminal random coil for the protein function we examined how small angle X-ray scattering of the wild-type S100A4 and its C-terminal deletion mutant (residues 1-88, Δ13) changes upon Ca2+ binding. We found that the scattering intensity of wild-type S100A4 changes substantially in the 0.15-0.25 Å-1 q-range whereas a similar change is not visible in the C-terminus deleted mutant. Ensemble optimization SAXS modeling indicates that the entire C-terminus is extended when Ca2+ is bound. Pulsed field gradient NMR measurements provide further support as the hydrodynamic radius in the wild-type protein increases upon Ca2+ binding while the radius of Δ13 mutant does not change. Molecular dynamics simulations provide a rational explanation of the structural transition: the positively charged C-terminal residues associate with the negatively charged residues of the Ca2+-free EF-hands and these interactions loosen up considerably upon Ca2+-binding. As a consequence the Δ13 mutant has increased Ca2+ affinity and is constantly loaded at Ca2+ concentration ranges typically present in cells. The activation of the entire C-terminal random coil may play a role in mediating interaction with selected partner proteins of S100A4.

MeSH terms

  • Algorithms
  • Calcium / chemistry*
  • Calorimetry
  • Crystallography, X-Ray
  • Humans
  • Molecular Dynamics Simulation
  • Mutation*
  • Neoplasm Metastasis
  • Nonmuscle Myosin Type IIA / chemistry
  • Protein Binding
  • Protein Structure, Tertiary
  • S100 Calcium-Binding Protein A4
  • S100 Proteins / chemistry*
  • Scattering, Radiation
  • Thermodynamics

Substances

  • S100 Calcium-Binding Protein A4
  • S100 Proteins
  • S100A4 protein, human
  • Nonmuscle Myosin Type IIA
  • Calcium

Grant support

The work was supported by the Swedish Research Council, Harald and Greta Jeanssons Foundation, Hungarian Scientific Research Fund (OTKA K108437, NK81950, OTKANK101072). DIS and MVP acknowledge support from the German Ministry of Education andScience (BMBF) project BIOSCAT, contract 05K20912. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.