Protease nexin-2/amyloid beta-protein precursor regulates factor VIIa and the factor VIIa-tissue factor complex

Protease nexin-2/amyloid beta-protein precursor (PN-2/A beta PP) and its Ku nitz protease inhibitory (KPI) domain were characterized as inhibitors of f actor VIIa (FVIIa) and factor VIIa-tissue factor complex (FVIIa-TF). PN-2/A beta PP and KPI domain inhibited FVIIa with an apparent K-i of 1.1+/-0.2x 10(-7) M and 1.5+/-0.1x10(-7) M, respectively. When soluble tissue factor ( TF1-219) was present, there was increased FVIIa inhibition by PN-2/A beta P P or KPI domain (K-i=7.81+/-0.3x10(-8) M and 6.8+/-0.6x10(-8) M, respective ly). When relipidated tissue factor (TF1-243) was present, the K-i of FVIIa inhibition by PN-2/A beta PP increased 4.7-fold further (K-i=1.65x 10(-8) M). PN-2/A beta PP complexed with FVIIa, as shown on gel filtration and sol id phase binding assay. The apparent second-order rate constant of inhibiti on of FVIIa by PN-2/A beta PP in the absence of TF1-219 (k"=7.5+/-4.0x10(4) M(-1)min(-1)) was less than that of the FVIIa-TF1-219 complex (k"=1.6+/- 0 .2x10(5) M(-1)min(-1)). Antithrombin in the absence of TF1-219 also had a l ower apparent second-order rate constant of inhibition (k"=1.8+/-0.3x10(3) M(-1)min(-1)) than in its presence (k"=1.6+/-0.3x10(5) M(-1)min(-1)). In a mixture that included FVIIa, relipidated TF1-243 and factor X, PN-2/A beta PP or KPI domain had an IC50 at 65 and 250 nM, respectively; antithrombin a nd heparin (1 U/mL) had an IC50 of 12.8 nM. These data indicate that tissue factor promoted the inhibition of FVIIa by PN-2/A beta PP or KPI domain, b ut antithrombin was a better inhibitor of soluble FVIIa-TF in extrinsic ten ase. (C) 2000 Elsevier Science Ltd. All rights reserved.