Indirect Role of Ca²⁺ in the Assembly of Extracelluar Matrix Proteins

¹ University of Illinois at Urbana-Champaign

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

Submitted on January 9, 2008
Revised on January 30, 2008
Accepted on 29 February 2008

	Abstract

We have investigated through molecular dynamics the binding properties at the interface between the C-type lectin subdomain (CLD) of aggrecan and the fibronectin type III domains (^4-5FnIII) of tenasin, in particular the mechanistically unknown, but essential role of Ca²⁺ ions in the binding. The binding between the CLD and ^4-5FnIII is critical in cross-linking aggrecan, hyaluronan and tenascin to form extended protein networks found in tissues such as cartilage. None of the structurally resolved Ca²⁺ ions in the complex of the CLD and ^4-5FnIII is directly bridging the two proteins. However, one of the Ca²⁺ ions (Ca2) is found to play the role of maintaining the structure of the L4 loop at the CLD binding surface, thus facilitating a high affinity binding between the CLD and ^4-5FnIII. Removal of this Ca²⁺ ion causes a drastic structural change in the L4 loop, which presumably hinders the binding of the CLD to the ^4-5FnIII. The other bound Ca²⁺ ions (Ca1 and Ca3) have no significant effect on the structure of the CLD binding surface, and thus are not expected to affect the binding. Our results might also suggest that the role of Ca2 in maintaining the structure of the L4 loop is most important during the binding process. Once the complex is formed, the dependence of the complex on the structuring role of Ca2 is reduced. In response to tensile force, CLD and ^4-5FnIII separate by breaking first the electrostatic interactions at the interface, followed by the hydrophobic ones. The sequence of the unbinding events and the maximum force required to separate the two proteins are independent of the presence of the Ca²⁺ ions, underlying the indirect role of Ca²⁺ in the binding.

Key Words: calcium, extracellular matrix, fibronectin, mechanical proteins, molecular dynamics, protein assembly