The importance of vicinal cysteines, C1669 and C1670, for von Willebrand factor A2 domain function

Abstract
The von Willebrand factor (VWF) A2 crystal structure has revealed the presence of a rare vicinal disulfide bond between C1669 and C1670, predicted to influence domain unfolding required for proteolysis by ADAMTS13. We prepared VWF A2 domain fragments with (A2-VicCC, residues 1473-1670) and without the vicinal disulfide bond (A2-ΔCC, residues 1473-1668). Compared with A2-ΔCC, A2-VicCC exhibited impaired proteolysis and also reduced binding to ADAMTS13. Circular dichroism studies revealed that A2-VicCC was more resistant to thermal unfolding than A2-ΔCC. Mutagenesis of C1669/C1670 in full-length VWF resulted in markedly increased susceptibility to cleavage by ADAMTS13, confirming the important role of the paired vicinal cysteines in VWF A2 domain stabilization.