Kinetics of the inhibition of factor Xa and the tissue factor-factor VIIa complex by the tissue factor pathway inhibitor in the presence and absence of heparin.
The inhibition by tissue factor pathway inhibitor (TFPI) of its two target enzymes--factor Xa and the tissue factor-factor VIIa complex (TF:VIIa)--has been studied under near-physiological reactant concentrations and conditions. Over a TFPI range of 0-1 nM, the rate of inhibition of factor Xa, in the presence of Ca2+ and anionic phospholipid vesicles at 37 degrees C, was proportional to TFPI concentration, giving an association rate, k1, of 0.96 x 10(9) M-1 min-1. Factor Xa inhibition did not proceed to completion, the reaction attaining a near-equilibrium that was dependent on the TFPI concentration. The estimated dissociation rate of the TFPI:Xa complex, k-1, was independent of TFPI concentration, with a mean value of 0.02 min-1. The resulting calculated value of K1, the apparent dissociation constant for the initial step, is 21 pM. Slow decay of the remaining factor Xa in such incubations, detectable after attainment of the rapid initial near-equilibrium, confirmed the two-step mechanism proposed by Huang et al. (1993) [J. Biol. Chem. 268, 26950-26955], but did not permit determination of a rate constant for the second step. Omission of anionic phospholipid had no significant effect on either k1 or k-1. A high-molecular-weight fraction of heparin, at saturating levels (> or = 0.05 unit/mL, congruent to 25 nM), increased k1 2-fold, with no detectable effect on k-1. The second stage of TFPI action was studied by preformation of the TFPI:Xa complex, and its incubation with the TF:VIIa complex in the presence of factor X.(ABSTRACT TRUNCATED AT 250 WORDS)
Jesty
J
J
Department of Medicine, State University of New York, Stony Brook 11794-8151.
Wun
T C
TC
Lorenz
A
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D000595:Amino Acid Sequence; D015942:Factor VIIa / Q000037:antagonists & inhibitors*; D065427:Factor Xa Inhibitors; D006493:Heparin / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008074:Lipoproteins / Q000494:pharmacology*; D008956:Models, Chemical; D008969:Molecular Sequence Data; D013925:Thromboplastin / Q000037:antagonists & inhibitors* / Q000378:metabolism
D065427:Factor Xa Inhibitors; D008074:Lipoproteins; C051928:lipoprotein-associated coagulation inhibitor; D006493:Heparin; D013925:Thromboplastin; D015942:Factor VIIa