Cadherin Diffusion in Supported Lipid Bilayers Exhibits Calcium-Dependent Dynamic Heterogeneity

Ca²⁺ ions are critical to cadherin ectodomain rigidity, which is required for the activation of adhesive functions. Therefore, changes in Ca²⁺ concentration, both in vivo and in vitro, can affect cadherin conformation and function. We employed single-molecule tracking to measure the diffusion of cadherin ectodomains tethered to supported lipid bilayers at varying Ca²⁺ concentrations. At a relatively high Ca²⁺ concentration of 2 mM, cadherin molecules exhibited a fast diffusion coefficient that was identical to that of individual lipid molecules in the bilayer (D_fast ≈ 3 μm²/s). At lower Ca²⁺ concentrations, where cadherin molecules were less rigid, the ensemble-average cadherin diffusion coefficient was systematically smaller. Individual cadherin trajectories were temporally heterogeneous, exhibiting alternating periods of fast and slow diffusion; the periods of slow diffusion (D_slow ≈ 0.1 μm²/s) were more prevalent at lower Ca²⁺ concentration. These observations suggested that more flexible cadherin ectodomains at lower Ca²⁺ concentration alternated between upright and lying-down conformations, where the latter interacted with more lipid molecules and experienced greater viscous drag.