Cys-rich domain amino acid sequence and sites of mutagenesis. (A) Domain organization of ADAMTS13. The Cys-rich domain comprises amino acids Lys440-Cys555. (B) The amino acids sequence of the Cys-rich domain contains a region that is highly conserved among ADAMTS family members (highlighted in gray) and a nonconserved region (indicated). In the nonconserved region, charged amino acids were individually mutated to alanine (bold and larger font). Amino acids that were mutated to asparagine for the introduction of an NXT glycosylation motif are indicated (Gly1-6). See supplemental Figure 2 for the location of each variant on a model of ADAMTS13. (C) Cys-rich domain sequence swap variants were generated by substitution of selected sequences for the corresponding sequence in ADAMTS1. Variants named Cys1-6 were used for functional analysis. Those swap variants labeled # were generated and expressed, but not secreted.