Structural Insight into the Role of Thrombospondin-1 Binding to Calreticulin in Calreticulin-Induced Focal Adhesion Disassembly

摘要

Thrombospondin-1 (TSP1) binding to calreticulin (CRT) on the cell surface stimulates associationnof CRT with LDL receptor-related protein (LRP1) to signal focal adhesion disassembly and engagement ofncellular activities. The structural basis for this phenomenon is unknown. We studied the binding thermo-ndynamics of the TSP1-CRT complex and the conformational changes in CRT induced by binding to TSP1nwith combined binding free energy analysis, molecular dynamics simulation, and anisotropic network modelnrestrained molecular dynamics simulation. Results showed that mutations of Lys 24 and Lys 32 in TSP1 tonAla and of amino acids 24-26 and 32-34 in CRT to Ala significantly weakened the binding of TSP1 andnCRT,which is consistentwith experimental results.Upon validation of the calculated binding affinity changesnof the TSP1-CRT complex by mutations in key residues in TSP1 and CRT with the experimental results, wenperformed conformational analyses to understand the role of TSP1 binding to CRT in the induction ofnconformational changes in CRT. Conformational analyses showed that TSP1 binding to CRT resulted in anmore “open” conformation and a significant rotational change for the CRT N-domain with respect to thenCRT P-domain, which could expose the potential binding site(s) in CRT for binding to LRP1 to signal focalnadhesion disassembly. Results offer structural insight into the role of TSP1 binding to CRT in CRT-inducednfocal adhesion disassembly.