Kininogen cleavage by Plm. (A) Schematic diagrams of human HK (top) and LK (bottom) showing domain structure and the disulfide bond connecting the N and C termini. The position of the BK nanopeptide is shown in black. (B-G) Coomassie Blue–stained SDS polyacrylamide gels showing HK and LK cleavage. At indicated times, samples were removed into nonreducing sample buffer and size fractionated on 10% SDS–polyacrylamide gels, followed by staining with Coomassie Blue. (B) Time course of HK cleavage by PKa. Human plasma–derived HK (200 nM) was incubated with PKa (2 nM) in reaction buffer at 37°C. The right panel is a western blot of samples from a reaction similar to that in panel D using an antibody to BK. (C) Time course of LK cleavage by PKa. Human plasma–derived LK (200 nM) was incubated with 2 (left) or 50 nM (right) of PKa in reaction buffer at 37°C. (D-E) Human plasma–derived HK (800 nM) incubated with 160 nM of Plm-Lys³¹¹ (D) or Plm-Glu³¹¹ (E). Bottom shows western blots for samples from reactions in the top using an antibody to BK. (F-G) Human plasma–derived LK (200 nM) incubated with 50 nM of Plm-Lys³¹¹ (D) or Plm-Glu³¹¹ (E). For all panels, positions of molecular mass standards (kDa) are indicated on the left. Positions of standards for uncleaved HK or LK, cleaved forms of HK (Cl HK) or (Cl LK), and PKa or Plm are indicated on the right. Numbers at the tops of gels represent incubation times in minutes.