Protein S Thr103Asn mutation associated with type II deficiency reproduced in vitro and functionally characterised.

Abstract

Protein S functions as a cofactor to activated protein C (APC) in the degradation of FVa and FVIIIa. In protein S, the thrombin sensitive region (TSR) and the first EGF-like domain are important for expression of the APC cofactor activity. A naturally occurring Thr103Asn (T103N) mutation in the first EGF-like domain of protein S has been associated with functional (type II) protein S deficiency. To elucidate the functional consequences of the T103N mutation, recombinant protein S mutant was expressed in mammalian cells and functionally characterised. The expression level of protein S T103N from transiently transfected COS 1 cells was equal to that of wild type protein S. The mutant protein S and wild type protein S were also expressed in 293 cells after stable transfection, and the recombinant proteins purified. In APTT- and PT-based coagulation assays, the mutant protein demonstrated approximately 50% lower anticoagulant activity as compared to wild type protein S. The functional defect was further investigated in FVa- and FVIIIa-degradation assays. The functional defect of mutant protein S was attenuated at increasing concentrations of APC. The results demonstrate the region around residue 103 of protein S to be of functional importance, possibly through a direct interaction with APC.

• Authors: T K Giri, P García de Frutos, B Dahlbäck
• Journal: Thrombosis and haemostasis (Thromb Haemost) Vol. 84 Issue 3 Pg. 413-9 (Sep 2000) ISSN: 0340-6245 [Print] Germany
• PMID: 11019964 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Protein C
• Protein S
• Reagent Kits, Diagnostic
• Recombinant Proteins
• factor V Leiden
• Factor Va
• Factor VIIIa
• Factor V

Topics

• Amino Acid Substitution
• Animals
• Blood Coagulation Tests
• COS Cells
• Dose-Response Relationship, Drug
• Factor V (drug effects, metabolism)
• Factor VIIIa (drug effects, metabolism)
• Factor Va (drug effects, metabolism)
• Humans
• Immunoblotting
• Mutagenesis, Site-Directed
• Partial Thromboplastin Time
• Point Mutation
• Protein C (metabolism, pharmacology)
• Protein S (genetics, pharmacology)
• Protein S Deficiency (genetics)
• Reagent Kits, Diagnostic
• Recombinant Proteins (genetics)
• Transfection