Molecular interactions of the intrinsic activation complex of coagulation: binding of native and activated human factors IX and X to defined phospholipid vesicles.
The assembly of proteins of the intrinsic activation complex has been partially elucidated. In the present study we examine the association of gamma-carboxylated serine proteinase zymogens factors IX and X, and their proteolytically activated counterparts factors IXa and Xa to unilamellar lipid vesicles of defined composition using three types of physical measurement. Utilizing relative light scatter to estimate the dissociation constants for binding in the presence of calcium ions, it appears that factor IXa (0.93 +/- 0.37 microM) may preferentially associate with phospholipids relative to factor IX (0.35 +/- 0.08 microM). In contrast, factor X (0.34 +/- 0.14 microM), the substrate for factor IXa, appears to bind to phospholipid with a higher affinity than factor Xa (0.58 +/- 0.13 microM). These observations are compatible with the hypothesized dynamics where the forward 'traffic' is facilitated by favoring the association of factor IXa with factor X. The dissociation constants were estimated by molecular exclusion chromatography (1.1 - 2.5 microM) and do not reflect these relative and ordered differences in association with lipid vesicles. Quasi-elastic light scatter analyses indicate that each protein appears to saturate the same vesicle surface, consistent with competition for similar surface lipids, although the molecular shell formed by factor Xa (36 A) is smaller, suggesting that it has a different packing on the phospholipid surface than the other proteins (64-79 A). The pattern of preferential affinities for phospholipid is consistent with a kinetically functional forward traffic through the reaction precursors to products, and suggests that these preferential affinities may assist in the ordering of the four proteins in the intrinsic activation complex.
D002118:Calcium / Q000502:physiology; D002850:Chromatography, Gel; D005164:Factor IX / Q000378:metabolism*; D015949:Factor IXa; D005170:Factor X / Q000378:metabolism*; D015951:Factor Xa; D006801:Humans; D008027:Light; D008970:Molecular Weight; D010743:Phospholipids / Q000378:metabolism*; D011485:Protein Binding; D012542:Scattering, Radiation
D010743:Phospholipids; D005164:Factor IX; D005170:Factor X; D015949:Factor IXa; D015951:Factor Xa; D002118:Calcium