Structural effects of the 17G2 mAb binding on the CUB1-2 interface. (A) Differential HDX heat map in the rainbow color scale (blue, high –ΔHDX; green, low ΔHDX; red, high +ΔHDX) plotted onto the ADAMTS13 CUB1-2 crystal structure with insets highlighting CUB1-2 interface residues that became less (Val¹²⁷⁰, Tyr¹²⁹⁶, and Phe¹³²⁸) or more (Arg¹²⁴⁷, Gln¹³⁰², Phe¹³⁰⁴, and Gly¹³⁰⁵) mobile upon 17G2 mAb binding. Our mAb 1C4 recognizes a conformationally sensitive epitope in the spacer domain of open ADAMTS13. All control CUB1-2 mutants (green) showed a similarly closed conformation as WT ADAMTS13. (B) Despite its lowered mobility upon 17G2 binding, the conformation of the Y1296P interface mutant (blue) remained similarly closed as for WT ADAMTS13, whereas its conformation could still be opened by preincubation of the 17G2 mAb.