Recombinant FXII. (A) Schematic diagrams of FXII, αFXIIa, and βFXIIa showing noncatalytic (white boxes) and protease (pink boxes) domains. Positions of the active site serine (Ser⁵⁴⁴) are indicated by black bars. Sites of proteolysis during activation are indicated by arrows. Cleavage after Arg³⁵³ (black arrow) converts FXII to αFXIIa. Cleavage of αFXIIa after Arg³³⁴ (white arrow) separates the noncatalytic and protease domains, forming βFXIIa. FXII noncatalytic domains are the fibronectin type 2 (F2), epidermal growth factor (EGF), fibronectin type 1 (F1), and kringle (K) domains, and a proline-rich region (PRR). (B) Schematic diagrams of FXII with lysine or arginine replacement of Thr³⁰⁹ (gray arrow). Cleavage after Lys/Arg³⁰⁹ creates 2 proteins, δHC and δFXII. (C) Nonreducing SDS-PAGE of purified FXII (∼3 μg per lane). Shown are FXII-WT, FXII-Lys³⁰⁹, FXII-Arg³⁰⁹, and FXII-3C. Reducing (D) and nonreducing (E) SDS-PAGE of the FXIIa (activated) forms of FXII shown in panel C. Proteins were activated by incubation with dextran sulfate. Positions of molecular mass standards in kilodaltons are shown to the left of the images in panels C-E. To the right of these images are markers for FXII, activated FXII (αFXIIa), HC of αFXIIa (HC), LC of αFXIIa (LC), HC of FXII-Lys³⁰⁹ or FXII-Arg³⁰⁹ cleaved after residue 309 (δHC), and FXII residues Thr³¹⁰ to Ser⁵⁹⁶ (δFXII).