Protein C inhibitor. Purification from human plasma and characterization.
Protein C inhibitor was isolated from human plasma using conventional chromatographic technique consisting of barium citrate adsorption, polyethylene glycol fractionation, DEAE-Sepharose CL-6B treatment, ammonium sulfate fractionation, dextran sulfate-agarose chromatography, gel filtration on ACA-44, and DEAE-Sephacel chromatography. The purified protein C inhibitor is a single polypeptide chain with an apparent Mr = 57,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The inhibitor is heterogeneous in pI: six pIs exist between pH 7.4 and 8.6. The inhibitor was shown to be different from the already known plasma protease inhibitors by chemical and immunological analyses. It migrates to the late alpha 1-globulin region on agarose gel electrophoresis. The inhibitor reduced the amidolytic activity of activated protein C noncompetitively by forming a 1:1 molar complex with the enzyme, determined by the use of a fluorogenic substrate toward activated protein C (Boc-Leu-Ser-Thr-Arg-4-methylcoumaryl-7-amide). The inhibition constant (Ki) of the inhibitor against activated protein C was 5.8 x 10(-8) M. The inhibitor also blocked the prolongation of activated partial thromboplastin time by activated protein C. The immunoglobulin which was produced by the inhibitor completely removed the inhibitory activity present in normal human plasma against activated protein C. This suggests that the inhibitor which we have isolated is the only inhibitor in plasma against activated protein C.
D000596:Amino Acids / Q000032:analysis; D001779:Blood Coagulation Factors / Q000378:metabolism; D001798:Blood Proteins / Q000302:isolation & purification* / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D006023:Glycoproteins / Q000378:metabolism; D006801:Humans; D005779:Immunodiffusion; D007122:Immunoelectrophoresis; D007700:Kinetics; D008970:Molecular Weight; D011486:Protein C; D018046:Protein C Inhibitor
D000596:Amino Acids; D001779:Blood Coagulation Factors; D001798:Blood Proteins; D006023:Glycoproteins; D011486:Protein C; D018046:Protein C Inhibitor