title: Lateral self-assembly of E-cadherin directed by cooperative calcium binding.
abstract: We report the Ca2+ binding characteristics of recombinant Ecad12, a construct spanning the first two repeats of epithelial cadherin, and demonstrate the links between Ca2+ binding and dimer formation. Sedimentation equilibrium and dynamic light scattering experiments show that weak dimerization of Ecad12 occurs in the presence of 10 mM Ca2+ (KdP = 0.17 mM), while no appreciable dimer formation was detected in the absence of Ca2+. Ca2+-induced dimerization was also observed in electron microscopy images of Ecad12. We conclude from Ca2+ titration experiments monitored by tryptophan fluorescence and flow dialysis that dimerization does not affect the equilibrium binding constant for Ca2+. However, the value of the Hill coefficient for Ca2+ binding increases from 1.5 to 2.4 as the protein concentration increases, showing that dimer formation largely contributes to the cooperativity in Ca2+ binding. Based on these observations and previous crystallographic studies, we propose that calcium acts more likely as a geometrical aligner ensuring the proper assembly of cadherin molecules, rather than a simple adhesive.
authors.affiliation: Ontario Cancer Institute and Department of Medical Biophysics, University of Toronto, Canada.
authors.forename: J R
authors.initials: JR
authors.lastname: Alattia
authors.forename: J B
authors.initials: JB
authors.lastname: Ames
authors.forename: T
authors.initials: T
authors.lastname: Porumb
authors.forename: K I
authors.initials: KI
authors.lastname: Tong
authors.forename: Y M
authors.initials: YM
authors.lastname: Heng
authors.forename: P
authors.initials: P
authors.lastname: Ottensmeyer
authors.forename: C M
authors.initials: CM
authors.lastname: Kay
authors.forename: M
authors.initials: M
authors.lastname: Ikura
mesh_terms: D001665:Binding Sites; D015820:Cadherins / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D002118:Calcium / Q000378:metabolism*; D019281:Dimerization; D004848:Epithelium / Q000378:metabolism; D008027:Light; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008958:Models, Molecular; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism / Q000648:ultrastructure; D012542:Scattering, Radiation
chemical_list: D015820:Cadherins; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D002118:Calcium