Inhibition of tissue factor-factor VIIa-catalyzed factor X activation by factor Xa-tissue factor pathway inhibitor - A rotating disc study on the effect of phospholipid membrane composition

The physiological inhibitor of tissue factor (TF) factor Wa (FVIIa), full-l ength tissue factor pathway inhibitor (TFPIFL) in complex with factor Xa (F Xa), has a high affinity for anionic phospholipid membranes, The role of an ionic phospholipids in the inhibition of TF.FVIIa-catalyzed FX activation w as investigated. FXa generation at a rotating disc coated with TF embedded in a membrane composed of pure phosphatidylcholine (TF PC) or 25% phosphati dylserine and 75% phosphatidylcholine (TF.PSPC) was measured in the presenc e of preformed complexes of FXa.TFPIFL or FXa.TFPI1-161 (TFPI lacking the t hird Kunitz domain and C terminus). At TF.PC, FXa.TFPIFL and FXa.TFPI1-161 showed similar rate constants of inhibition (0.07 x 10(8) M-1 s(-1) and 0.1 x 10s RI-1 S-1, respectively). With phosphatidylserine present, the rate c onstant of inhibition for FXa.TFPIFL increased 3-fold compared with a 9-fol d increase in the rate constant for FXa.TFPI1-161. Incubation of TF.PSPC wi th FXa TFPI,, in the absence of FVIIa followed by depletion of solution FXa .TFPI1-161 showed that FXa.TFPIFL remained bound at the membrane and pursue d its inhibitory activity. This was not observed with FXa.TFPI1-161 or at T F.PC membranes. These data suggest that the membrane-bound pool of FXa. TFP Ia may be of physiological importance in an on-site regulation of TF.FVIIa activity.