Secretion of PP/Gla domain-chimeric PZ and FX. Construction of the chimeric proteins is illustrated on the top: domains in PZ and FX are shown as gray and white boxes, respectively. In PZ_PPmut, PZ was mutated at the cleavage site of its PP. Twenty micrograms expression vector for each construct were transfected to HEK 293 cells, and the cells were incubated with warfarin (W) or Vit.K (K). Barium citrate adsorption was carried out from 0.5 mL medium, and the precipitate was dissolved in a 50-μL 0.1 M EDTA solution. The medium, barium citrate adsorbate, and cell lysates were analyzed by Western blot analysis using an anti-PZ (A) or anti-FX (B) antibody.