A protease isolated from human plasma activating factor VII independent of tissue factor.
An activity detected in a prothrombin complex concentrate, termed 'thrombin-like' due to its amidolytic properties, was recently reported by another working group. This serine-protease revealed partial structural homology with a 'hepatocyte growth factor activator'. An impact of this protease on coagulation has not yet been described. The protease was isolated from plasma fractions by ion exchange chromatography and adsorption to immobilized heparin and/or aprotinin. Clotting tests including the FVIIa-rTF assay were performed employing coagulometry. A monoclonal antibody-derived F(ab')2 to FVIII was used to investigate the FVIII bypassing activity (FEIBA). The identity of the-protease with the so-called 'thrombin-like' protease was supported by sequencing of the amino-termini. Its amidolytic activity was significantly enhanced in the presence of calcium and/or heparin. Incubation with purified FVII revealed the generation of FVIIa, but was prevented by pre-incubation of the protease with aprotinin. In contrast, purified FV and FVIII were inactivated. Studying coagulation parameters, clotting times like plasma recalcification times and the prothrombin times were found to be shortened by addition of the protease. Employing a FVIII-inhibitory F(ab')2 and enhancing clotting times significantly, FEIBA of the protease was found. We demonstrated that the isolated protease activates FVII independent of tissue factor. Net acceleration of coagulation was found in several global clotting assays resulting in an in vitro FEIBA. The physiological relevance of these findings deserves further investigation.
Römisch
J
J
Centeon Pharma GmbH, Research, Marburg, Germany. Roemisch@msmbwmd.hoechst.com
Feussner
A
A
Vermöhlen
S
S
Stöhr
H A
HA
D000581:Amidohydrolases / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism; D007611:Aprotinin / Q000378:metabolism; D001777:Blood Coagulation / Q000187:drug effects; D002413:Cations, Divalent / Q000494:pharmacology; D010450:Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D005167:Factor VII / Q000378:metabolism*; D005169:Factor VIII / Q000378:metabolism; D015942:Factor VIIa / Q000096:biosynthesis; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D020539:Sequence Analysis, Protein; D013925:Thromboplastin; D014914:Whole Blood Coagulation Time
D002413:Cations, Divalent; D005167:Factor VII; D005169:Factor VIII; D006493:Heparin; D013925:Thromboplastin; D007611:Aprotinin; D010450:Endopeptidases; D015942:Factor VIIa; D000581:Amidohydrolases