Modulation of E-cadherin monomer folding by cooperative binding of calcium ions.
Classical cadherins are transmembrane glycoproteins involved in calcium-dependent cell-cell adhesion. Calcium ions are coordinated at the interface between successive modules of the cadherin ectodomain and are thought to regulate the adhesive interactions of cadherins when present at millimolar concentrations. It is widely accepted that calcium plays a critical role in cadherin-mediated cell-cell adhesion, but the nature of cadherin-calcium binding remains a matter of debate. We investigated the parameters of noncovalent cadherin-calcium binding, using the two N-terminal modules of E-cadherin (E/EC12) with a native N-terminal end and nondenaturing electrospray ionization mass spectrometry. By directly visualizing the molecular complexes, we demonstrated that E/EC12 binds three calcium ions, with an average KD of 20 +/- 0.7 microM. These calcium ions bound cooperatively to E/EC12 in its monomeric state, and these properties were not modified by an N-terminal extension consisting of a single methionine residue. This binding induced specific structural changes, as shown by assessments of protease sensitivity, circular dichroism, and mass spectrometry. Furthermore, the D103A mutation (a residue involved in E-cadherin adhesive function) modified calcium binding and led to a loss of cooperativity and the absence of structural changes, despite calcium binding. As the amino acids involved in calcium binding are found within the cadherin consensus motif, our findings may be relevant to other members of the cadherin family.
Courjean
Olivier
O
Morphogenèse cellulaire et progression tumorale, Institut Curie, CNRS UMR 144, Paris, France.
Chevreux
Guillaume
G
Perret
Emilie
E
Morel
Anne
A
Sanglier
Sarah
S
Potier
Noelle
N
Engel
Jürgen
J
van Dorsselaer
Alain
A
Feracci
Hélène
H
D000494:Allosteric Regulation; D015820:Cadherins / Q000737:chemistry* / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D007477:Ions / Q000378:metabolism; D008954:Models, Biological; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D015820:Cadherins; D007477:Ions; D050505:Mutant Proteins; D011994:Recombinant Proteins; D002118:Calcium