In vitro interaction of C1-inhibitor with thrombin

Massimo Cugno, I. Bos, Y. Lubbers, C. Erik Hack, A. Agostoni

Research output: Contribution to journalArticle

27 Citations (Scopus)

Abstract

Previous observations of increased generation of thrombin during acute attacks of angioedema in plasma of patients with C1-inhibitor (C1-INH) deficiency prompted us to evaluate the interaction of C1-INH with thrombin in both purified systems and human plasma. For this purpose, we used several methods: (1) sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting analysis; (2) enzyme-linked immunosorbent assays to measure complexes between C1-INH and thrombin and inactivated C1-INH; and (3) kinetic studies using a chromogenic assay. We found that the interaction of purified C1-INH with thrombin is associated with the formation of bimolecular complexes of molecular weight (Mr) 130 000 and 120 000 as well as with the appearance of a cleaved form of C1-INH of Mr 97 000. The kinetic studies of inhibition of thrombin by C1-INH showed an average second-order rate constant of 19/s per mol/l, which was significantly increased in the presence of heparin. The addition of thrombin to human plasma was not associated with detectable C1-INH-thrombin complex formation or with cleavage of C1-INH. In conclusion, our data demonstrate that C1-INH upon interaction with thrombin, in part, forms enzyme-inhibitor complexes and, in part, is cleaved. The low second-order rate constant and the lack of a significant interaction in plasma suggest that the inhibition of thrombin by C1-INH has a minor role in circulating blood; however, its role might be important at the endothelial surface, where high concentrations of glycosaminoglycans occur.

Original languageEnglish
Pages (from-to)253-260
Number of pages8
JournalBlood Coagulation and Fibrinolysis
Volume12
Issue number4
DOIs
Publication statusPublished - 2001

Fingerprint

Thrombin
Hereditary Angioedemas
In Vitro Techniques
Angioedema
Enzyme Inhibitors
Glycosaminoglycans
Immunoblotting
Sodium Dodecyl Sulfate
Heparin
Polyacrylamide Gel Electrophoresis
Molecular Weight
Enzyme-Linked Immunosorbent Assay

Keywords

  • C1-inhibitor
  • Complement
  • Contact system
  • Heparin
  • Thrombin

ASJC Scopus subject areas

  • Hematology

Cite this

In vitro interaction of C1-inhibitor with thrombin. / Cugno, Massimo; Bos, I.; Lubbers, Y.; Hack, C. Erik; Agostoni, A.

In: Blood Coagulation and Fibrinolysis, Vol. 12, No. 4, 2001, p. 253-260.

Research output: Contribution to journalArticle

Cugno, Massimo ; Bos, I. ; Lubbers, Y. ; Hack, C. Erik ; Agostoni, A. / In vitro interaction of C1-inhibitor with thrombin. In: Blood Coagulation and Fibrinolysis. 2001 ; Vol. 12, No. 4. pp. 253-260.
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