Cadherin mechanics and complexation: the importance of calcium binding

Abstract

E-cadherins belong to a family of membrane-bound, cellular adhesion proteins. Their adhesive properties mainly involve the two N-terminal extracellular domains (EC1 and EC2). The junctions between these domains are characterized by calcium ion binding sites, and calcium ions are essential for the correct functioning of E-cadherins. Calcium is believed to rigidify the extracellular portion of the protein, which, when complexed, adopts a rod-like conformation. Here, we use molecular dynamics simulations to investigate the dynamics of the EC1-2 portion of E-cadherin in the presence and in the absence of calcium ions. These simulations confirm that apo-cadherin shows much higher conformational flexibility on a nanosecond timescale than the calcium-bound form. It is also shown that although the apo-cadherin fragment can spontaneously complex potassium, these monovalent ions are incapable of rigidifying the interdomain junctions. In contrast, removal of the most solvent-exposed calcium ion at the EC1-2 junction does not significantly perturb the dynamical behavior of the fragment. We have also extended this study to the cis-dimer formed from two EC1-2 fragments, potentially involved in cellular adhesion. Here again, it is shown that the presence of calcium is an important factor in both rigidifying and stabilizing the complex.

FIGURE 1

Ribbon diagrams of EC1-2 fragments of E-cadherin. (a) Monomer, showing the three calcium ions (in green) bound at the domain junction and the axes used to characterize the relative position of the two domains. (b) cis-dimer of EC1-2 (PDB file 1FF5, (11)). The images in Figs. 1 and 4 were prepared using VMD (33).

FIGURE 2

Geometry of the interdomain calcium binding sites. The histogram at the bottom of the figure shows the evolution of the calcium binding sites in terms of the distance between the chelating atoms of axially opposed amino acid side chains: 1FF5 crystallographic values (solid), C3 (open), C2 (shaded), and C0 (crosshatched). For the values coming from EC1-2 monomer dynamics simulations, the standard deviation of each distance is indicated by error bars.

FIGURE 3

Occupation of the calcium binding sites during the dynamics simulations (time in picoseconds). Solid zones correspond to calcium binding, and shaded zones to potassium binding. Each group of three bars corresponds to the sites CA1, CA2, and CA3 reading from bottom to top.

FIGURE 4

RMSD with respect to the crystallographic conformation of the EC1-2 monomer. (a) Simulations C3 (thin shaded), C2 (thin solid), C0 (thick solid), and K3 (thick shaded). The break in the C2 line corresponds to restarting the simulation after removing a potassium ion from site CA3. (b) Values for EC1 (shaded) and EC2 (thick solid) during the C0 simulation. The thin solid line corresponds to domain EC2, excluding the loop residues 130–145.

FIGURE 5

Interdomain (a) bending angle and (b) torsion for EC1-2 during the C0 (solid line) and C3 simulations (shaded line).

FIGURE 6

Snapshots extracted at regular intervals from the C0 simulation showing interdomain flexibility in the absence of bound calcium ions. The axes of the domains are shown in black. The upper (EC1) domains of each snapshot have been placed in identical orientations.

FIGURE 7

RMSD with respect to the crystallographic cis-dimer structure of EC1-2 (shaded lines) during simulations D6 (a) and D0 (b). Solid lines show the RMSD fluctuations of each EC1-2 fragment with respect to their crystallographic conformation.

FIGURE 8

Interdomain (a) bending angle and (b) torsion for EC1-2 during the D6 (shaded line) and D0 simulations (solid line).

FIGURE 9

Evolution of the interface area during the simulations D6 (shaded) and D0 (solid).