Submitted March 24, 2006
Accepted July 24, 2006
A single high-affinity binding site for von Willebrand
Factor in collagen III, identified using synthetic
triple-helical peptides
Ton Lisman, Nicolas Raynal, Dafna Groeneveld, Ben Maddox, Anthony R Peachey, Eric G Huizinga, Philip G de Groot, and Richard W Farndale*
Department of Clinical Chemistry and Haematology, University Medical Centre Utrecht, Netherlands
Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom
Department of Crystal and Structural Chemistry, Utrecht University, Utrecht, Netherlands
* Corresponding author; email: rwf10{at}cam.ac.uk.
The essential event in platelet adhesion to the injured blood vessel wall is the binding to sub-endothelial collagen of plasma von Willebrand factor (VWF), a protein which interacts transiently with platelet glycoprotein (GP) Ib
1-3, slowing circulating platelets to facilitate firm adhesion through collagen receptors, including integrin 2
1 and GPVI. To locate the site in collagen that binds VWF, we synthesized 57 overlapping triple-helical peptides comprising the whole triple-helical domain of collagen III. Peptide #23 alone bound VWF, with similar affinity to that of native collagen III. Immobilized peptide #23 supported platelet adhesion under static and flow conditions, processes blocked by an antibody which prevents collagen from binding the VWF A3 domain. Truncated and alanine-substituted peptides derived from #23 either strongly interacted with both VWF and platelets, or lacked both VWF and platelet binding. Thus, we identified the sequence RGQOGVMGF (O is hydroxyproline) as the minimal VWF-binding sequence in collagen III.
