Alkylation of the cysteine thiol in E3CaG-1 ablates the VWF-reducing activity.

(A) E3CaG-1 or NEM-alkylated E3CaG-1 (NEM-E3CaG-1) was incubated with MPB, resolved on SDS-PAGE, and blotted with streptavidin-peroxidase to detect the biotin label. The positions of M_r markers are shown at left. (B) VWF (8 nM) was incubated with 80 nM TSP-1, 8 or 80 nM E3CaG-1 or 80 nM NEM-E3CaG-1. Aliquots of the reactions were analyzed for the average VWF multimer size (■) and for the generation of new thiols in VWF (▪). The dotted lines represent no change in VWF multimer size (top line) or thiols in VWF (bottom line). The asterisks indicate significant difference compared to control (P < .05).