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Partial rescue of naturally occurring active site factor X variants through decreased inhibition by tissue factor pathway inhibitor and antithrombin
Author(s) -
Ahnström Josefin,
Gierula Magdalena,
Temenu Joseph,
Laffan Michael A.,
Lane David A.
Publication year - 2020
Publication title -
journal of thrombosis and haemostasis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.947
H-Index - 178
eISSN - 1538-7836
pISSN - 1538-7933
DOI - 10.1111/jth.14627
Subject(s) - prothrombinase , tissue factor , thrombin , serine protease , tissue factor pathway inhibitor , chemistry , antithrombin , protease , factor x , thromboplastin , recombinant dna , microbiology and biotechnology , factor ixa , coagulation , biochemistry , enzyme , biology , platelet , medicine , immunology , heparin , gene
Abstract Background Activated coagulation factor X ( FX a) is the serine protease component of prothrombinase, the physiological activator of prothrombin. Factor X Nottingham (A404T) and Taunton (R405G) are two naturally occurring mutations, identified in families with a bleeding phenotype. Objective To characterize these FX variants functionally. Methods The activity and inhibition of recombinant FX variants were quantified in plasma‐based and pure component assays. Results The prothrombin times in FX ‐depleted plasma supplemented with FX Nottingham and Taunton were greatly increased compared to that of wild‐type ( WT ) FX . Kinetic investigations of activated variants in the prothrombinase complex showed k cat / K m reduced ~50‐fold and ~5‐fold, respectively, explaining the prolonged prothrombin time ( PT) . The substituted residues are located in the protease domain Na + ‐binding loop, important for the activity of FX a, as well as its inhibition. Both FX a Nottingham and Taunton showed reduced affinity for Na + . Plasma‐based thrombin generation assays triggered with 1 pmol/L tissue factor ( TF ) demonstrated only small differences in activities compared to WT FX , but large reductions at 10 pmol/L TF . Severely reduced inhibition of both FX a Nottingham and Taunton by tissue factor pathway inhibitor ( TFPI ) and antithrombin ( AT ), was shown in pure‐component FX a inhibition assays. Factor X a Nottingham and Taunton produced higher amounts of thrombin than WT FX a in pure‐component prothrombinase assays in the presence of TFPI and AT , explaining the results from the plasma‐based assay. Conclusions Factor X Nottingham and Taunton both display decreased proteolytic activity. However, their reduced activity in plasma triggered by low TF can be rescued by decreased inhibition by the natural FX a inhibitors, TFPI and AT .