Roles of the membrane-interactive regions of factor VIIa and tissue factor. The factor VIIa Gla domain is dispensable for binding to tissue factor but important for activation of factor X.
The roles of the putative membrane-interactive regions of factor VIIa (fVIIa) and tissue factor (TF) have been examined. Enzymatic removal of the 4-carboxyglutamic acid (Gla) domain of fVIIa had no effect on hydrolysis of a tripeptidyl chromogenic substrate in the absence or presence of TF. Additionally, Gla-domainless fVIIa (GdVIIa) was similar to native fVIIa in activating factor X in the absence of TF and phospholipid. However, GdVIIa in complex with recombinant soluble TF (sTF) was 76-fold less efficient in factor X activation than was fVIIa.sTF. The difference increased to 740-fold using TF relipidated in vesicles composed of 80% phosphatidylcholine and 20% phosphatidylserine (TF/PCPS). While Gla domain deletion produced a 10(3)-fold increase in the Kd for binding to TF/PCPS, the Kd for binding to TF/PC increased only 20-fold, and that for sTF in the absence of phospholipid increased 10-fold. Kd values for GdVIIa binding to TF/PCPS, TF/PC, or sTF were nearly identical. Thus, most of the binding energy required for formation of the fVIIa.TF complex was present even after Gla domain deletion. Both fVIIa and GdVIIa were capable of binding sTF in the presence of excess divalent metal-ion chelator, suggesting Ca(2+)-independent binding or the presence of a novel very high affinity Ca2+ binding site in fVIIa. The results demonstrate that the effect of the Gla domain on the Kd is apparent only in the presence of PS, and that interactions involving the fVIIa Gla domain and phospholipid are critical for efficient proteolysis of factor X on a membrane surface.
Neuenschwander
P F
PF
Oklahoma Medical Research Foundation, Oklahoma City 73104.
Morrissey
J H
JH
D015055:1-Carboxyglutamic Acid / Q000378:metabolism*; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D002118:Calcium / Q000494:pharmacology; D005164:Factor IX / Q000302:isolation & purification / Q000378:metabolism; D015942:Factor VIIa / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008081:Liposomes; D008433:Mathematics; D008962:Models, Theoretical; D010713:Phosphatidylcholines / Q000494:pharmacology; D010718:Phosphatidylserines / Q000494:pharmacology; D013925:Thromboplastin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
D008081:Liposomes; D010713:Phosphatidylcholines; D010718:Phosphatidylserines; D015055:1-Carboxyglutamic Acid; D005164:Factor IX; D013925:Thromboplastin; D015942:Factor VIIa; D002118:Calcium