Functional analysis of ADAMTS13 Cys-rich domain sequence swap variants. (A) A total of 5 nM ADAMTS13 or ADAMTS13 Cys1 variant in conditioned medium was incubated with 6 µM VWF115. Aliquots were taken and stopped with EDTA at 0, 30, and 90 minutes and analyzed by SDS-PAGE and Coomassie staining. Uncleaved VWF115 and cleavage products arising from specific ADAMTS13-mediated proteolysis are marked by arrows. (B) ADAMTS13 variants Cys2 to Cys4 were analyzed as in panel A, except that 2 nM ADAMTS13 was used and aliquots were taken at 0, 45, and 120 minutes. (C) ADAMTS13 variants Cys5 and Cys6 were analyzed as in panel B.