Lifetime Measurements Reveal Kinetic Differences between Homophilic Cadherin Bonds

¹ University of Illinois
² University of British Columbia
³ University of Vancouver

^* To whom correspondence should be addressed. E-mail: leckband{at}uiuc.edu.

Submitted on June 28, 2005
Revised on September 27, 2005
Accepted on 11 October 2005

	Abstract

Cadherins are multi-domain adhesion proteins whose interactions direct cell sorting during histogenesis. They determine cell adhesion specificity, but prior studies failed to identify physical differences that could underlie cell sorting. These single molecule studies identify kinetic and strength differences between different cadherins. They further demonstrate that the modular extracellular architecture of cleavage stage C-cadherin supports a multi-state binding mechanism. These multiple bonds exhibit a kinetic hierarchy of strengths that map to the different cadherin domains. The outer two N-terminal domains of C-cadherin form two bound states with dissociation rates of 3.9 and 0.02s-1. The latter is 25-fold slower than between the corresponding epithelial cadherin segments. In addition to the two fast bonds, the five-domain fragment (CEC1-5) forms two additional stronger, longer-lived bonds with dissociation rates of 0.00039 and 0.00001 s-1. We further quantified the lifetimes of bonds subject to a constant force, and thus identified multiple dissociation events with rates that agree quantitatively with the force spectroscopy data. The qualitative features are similar to those reported for epithelial cadherin. However, the significant differences in the dissociation rates of the outer domains, which include the specificity-determining region, suggest that kinetic differences may determine cadherin discrimination, rather than adhesion energies.

Key Words: Cadherin, biomembrane force probe, cell sorting, force spectroscopy, kinetics

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