Factor Xa Cleavage of Tissue Factor Pathway Inhibitor Is Associated with Loss of Anticoagulant Activity^[*]

Summary

Tissue factor : factor VIIa induced activation of blood coagulation is inhibited by the complex between factor Xa and tissue factor pathway inhibitor (factor Xa : TFPI). We recently reported that phospholipid-bound factor Xa reduces the high binding affinity of factor Xa : TFPI for negatively charged phospholipids by a partial degradation of TFPI (17). The present study was undertaken to elucidate the factor Xa cleavage sites in TFPI and to delineate the consequences of this proteolysis with respect to the inhibitory activity of factor Xa : TFPI. We found that phospholipid-bound factor Xa cleaves in TFPI the peptide bonds between Lys⁸⁶-Thr⁸⁷ and Arg¹⁹⁹-Ala²⁰⁰. Interestingly, Arg¹⁹⁹ is the P1 residue of the third Kunitz-type protease inhibitor domain. The fast cleavage of the Arg¹⁹⁹-Ala²⁰⁰ bond results in a 50-70% reduction of the anticoagulant activity of factor Xa : TFPI, as determined with a dilute tissue factor assay, but is not associated with a diminished inhibitory activity of factor Xa : TFPI towards TF : factor VIIa catalyzed activation of factor X. On the other hand, the slower cleavage of the Lys⁸⁶-Thr⁸⁷ peptide bond was associated with both a diminished anticoagulant and anti-TF : factor VIIa activity. Dissociation of factor Xa from the cleaved TFPI was not observed. These data provide evidence for a dual role of factor Xa since it is the essential cofactor in the TFPI-controlled regulation of TF-dependent coagulation as well as a catalyst of the inactivation of TFPI.

^* Supported by Grant 902-26-154 from the Dutch Organization for Scientific Research (NWO)

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