Role of gamma-carboxyglutamic acid residues in the binding of factor IXa to platelets and in factor-X activation.
To study the requirements for factor-IXa binding to platelets and factor-X activation, we examined the consequences of chemical modification (factor IXMOD) or enzymatic removal (factor IXDES) of gamma-carboxyglutamic acid (Gla) residues. In the presence of factor VIIIa and factor X, there were 344 (+/- 52) binding sites/platelet for factor IXaMOD (apparent dissociation constant [kdapp] = 4.5 +/- 0.9 nmol/L) and 275 (+/- 35) sites/platelet for factor IXaDES (kdapp = 5.0 +/- 0.8 nmol/L) compared with 580 (+/-65) sites/platelet for normal factor IXa (factor IXaN) (kdapp = 0.61 +/- 0.1 nmol/L) and 300 (+/-62) sites/platelet for factor IX (kdapp = 2.9 +/- 0.29 nmol/L). The concentrations of factor IXaN, factor IXaMOD and factor IXaDES required for half-maximal rates of factor-Xa formation were 0.67 nmol/L, 3.5 nmol/L, and 6.7 nmol/L. Whereas maximal velocities (Vmax) of factor Xa formation by factor IXaMOD (approximately 0.8 nmol/L.min-1) and factor IXaN (approximately 10.5 nmol/L.min-1), turnover numbers (kcat expressed as moles of factor Xa formed per minute per mole of factor IXa bound), and values of catalytic efficiency (kcat/Km) were normal, indicating that the decreased rates of factor X activation observed with factor IXaMOD and factor IXaDES are solely a consequence of the abnormal binding of these proteins to thrombin-activated platelets in the presence of factor VIIIa and factor X. Thus, factor IXa binding to platelets is mediated in part, but not exclusively, by high-affinity Ca2+ binding sites in the Gla domain of factor IX.
Rawala-Sheikh
R
R
Department of Medicine, Temple University School of Medicine, Philadelphia, PA 19140.
Ahmad
S S
SS
Monroe
D M
DM
Roberts
H R
HR
Walsh
P N
PN
D015055:1-Carboxyglutamic Acid / Q000097:blood* / Q000737:chemistry; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D015949:Factor IXa / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D005170:Factor X / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D015539:Platelet Activation / Q000187:drug effects; D013329:Structure-Activity Relationship; D013917:Thrombin / Q000494:pharmacology
D015055:1-Carboxyglutamic Acid; D005170:Factor X; D015949:Factor IXa; D013917:Thrombin; D002118:Calcium