HIGH-AFFINITY, SPECIFIC FACTOR IXA BINDING TO PLATELETS IS MEDIATED IN PART BY RESIDUES-3-11

被引:31
作者
AHMAD, SS
RAWALASHEIKH, R
CHEUNG, WF
JAMESON, BA
STAFFORD, DW
WALSH, PN
机构
[1] TEMPLE UNIV,SCH MED,SOL SHERRY THROMBOSIS RES CTR,PHILADELPHIA,PA 19140
[2] TEMPLE UNIV,SCH MED,DEPT BIOCHEM,PHILADELPHIA,PA 19140
[3] TEMPLE UNIV,SCH MED,DEPT MED,PHILADELPHIA,PA 19140
[4] UNIV N CAROLINA,DEPT BIOL,CHAPEL HILL,NC 27599
[5] THOMAS JEFFERSON UNIV,JEFFERSON CANC INST,PHILADELPHIA,PA 19107
关键词
D O I
10.1021/bi00206a006
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
To identify the amino acids in the Gla domain that mediate factor IXa binding to human platelets, we have used chimeric molecules and point mutations in the Gla domain of recombinant factor IX, based on molecular modeling using the coordinates of the Gla domain of bovine prothrombin, which reveals two surface structures whose sequences differ among factor IX, factor X, and factor VII. Binding to thrombin-activated platelets of factor IXa in the presence of factor VIIIa (2 units/mL) and factor X (1.5 mu M) revealed a stoichiometry of similar to 550 sites per platelet with a K-d of similar to 0.65 nM compared with a K-d of similar to 2.5 nM in the absence of factor VIIIa and factor X. In contrast, mutations of factor IX to factor X residues at positions 4 and 5 or at positions 9, 10, and 11 resulted in decreases in the number of sites and affinity of factor IXa binding in the presence or absence of factor VIIIa and factor X. A chimera consisting of the Gla domain of factor VII with factor IX residues at positions 33, 34, 35, 39, and 40 displayed abnormal factor IXa binding and a decreased V-max and a normal K-m for factor X activation, and the replacement of amino acid residues 3-10 with those of factor IX restored normal binding and factor X activation kinetics to this chimeric protein. Our data indicate that the high-affinity, specific binding of factor IXa to activated platelets in the presence or absence of factor VIIIa and factor X is the major determinant of rates of factor X activation and that both factor IXa binding and factor X activation are mediated at least in part by amino acids exposed on the surface of the Gla domain within positions 3-11, possibly by residues 4, 5, 9, 10, and 11.
引用
收藏
页码:12048 / 12055
页数:8
相关论文
共 22 条
[1]   COMPONENTS AND ASSEMBLY OF THE FACTOR-X ACTIVATING COMPLEX [J].
AHMAD, SS ;
RAWALASHEIKH, R ;
WALSH, PN .
SEMINARS IN THROMBOSIS AND HEMOSTASIS, 1992, 18 (03) :311-323
[2]  
AHMAD SS, 1989, J BIOL CHEM, V264, P20012
[3]  
AHMAD SS, 1992, J BIOL CHEM, V267, P8571
[4]  
AHMAD SS, 1989, J BIOL CHEM, V264, P3244
[5]  
AHMAD SS, 1990, J BIOL CHEM, V265, P20907
[6]   PLATELET RECEPTOR MEDIATED FACTOR-X ACTIVATION BY FACTOR-IXA - HIGH-AFFINITY FACTOR-IXA RECEPTORS INDUCED BY FACTOR-VIII ARE DEFICIENT ON PLATELETS IN SCOTT SYNDROME [J].
AHMAD, SS ;
RAWALASHEIKH, R ;
ASHBY, B ;
WALSH, PN .
JOURNAL OF CLINICAL INVESTIGATION, 1989, 84 (03) :824-828
[7]  
AHMAD SS, 1991, THROMB HAEMOSTASIS, V65, P800
[8]  
CHEUNG WF, 1992, J BIOL CHEM, V267, P20529
[9]  
CHEUNG WF, 1991, J BIOL CHEM, V266, P8797
[10]  
DERIAN CK, 1989, J BIOL CHEM, V264, P6615