Proteolysis of protein C in pooled normal plasma and purified protein C by activated protein C (APC)
| Autor: | Houria I. Hassouna, Christopher Quinn |
|---|---|
| Rok vydání: | 2002 |
| Předmět: |
Serine Proteinase Inhibitors
Immunoblotting Biophysics Enzyme-Linked Immunosorbent Assay In Vitro Techniques Biochemistry Epitope Thrombin Protein C deficiency Zymogen Protein A/G medicine Humans Blood Coagulation Serine protease chemistry.chemical_classification biology Serine Endopeptidases Organic Chemistry Antibodies Monoclonal Anticoagulants medicine.disease Molecular biology Enzyme Activation Kinetics Enzyme chemistry biology.protein Indicators and Reagents Protein C medicine.drug |
| Zdroj: | Biophysical Chemistry. 95:109-124 |
| ISSN: | 0301-4622 |
| DOI: | 10.1016/s0301-4622(01)00267-8 |
| Popis: | Protein C is a vitamin-K dependent zymogen of the anti-coagulant serine protease activated protein C (APC). In this paper, we report four lines of evidence that APC can activate protein C in pooled normal plasma, and purified protein C. First, the addition of APC to protein C-deficient plasma supplemented with protein C produces a prolongation of the clotting time of plasma that is proportional to the amount of protein C. This behavior was observed with APC from the Chromogenix APC resistance kit (Dia Pharm, Franklin, OH, USA) and from APC derived from the thrombin activation of human protein C (Enzyme Research Laboratories, South Bend, IN, USA). Secondly, using immunoblotting after gel electrophoresis, the disappearance of epitopes for monoclonal antibodies that recognize protein C but not APC indicates a time course for the activation by APC of protein C in pooled normal plasma and protein C purified from plasma. Thirdly, the same time course for the disappearance of protein C specific epitope can be followed using ELISA. Finally, protein C can be activated by APC as indicated by the increase in APC specific synthetic substrate Tryp–Arg–Arg– p nitroaniline hydrolysis. Kinetic data indicate a value of 4.7±0.4 mM −1 s −1 for the activation of protein C by APC under physiological conditions and in the presence of calcium. These observations document that APC must function not only in the inactivation of activated factors V and VIII, but also in the activation of protein C. This additional action of APC may be important to consider more broadly because of APC in the treatment of sepsis. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect