Determinants of the formation and activity of factor V-phospholipid complexes. II. Molecular properties of the complexes.
The stability, buoyancy, and intrinsic activity for factor V-phospholipid complexes were investigated. The decay of factor V activity in the absence of phospholipid followed first order kinetics; however, in the presence of several phospholipids a biphasic decay curve was observed. The addition of phosphatidylethanolamine to factor V produced only a small loss of activity in the first 2 minutes but decreased the subsequent rate of inactivation fourfold. A PE-factor V complex with a low bouyant density was separated from uncomplexed factor V by sucrose density ultracentrifugation. The association constant for this complex was 5 X 10(6) M-1 with approximately 2 moles of factor V bound per mole of lipid micelle. The isolated complex was capable of increasing prothrombin conversion 10-fold without additional phospholipid. A still lighter complex increased the rate of prothrombin conversion 18-fold. Phosphatidyl serine produced a concentration-dependent loss of up to 95% of the factor V activity in the first 2 minutes. After ultracentrifugation on a sucrose density gradient, a PS-factor V complex of increased density was detected. This complex failed to accelerate prothrombin conversion in the intrinsic two-stage assay. Except at very high concentrations, phosphatidylcholine did not alter the kinetics of inactivation of factor V. A factor V-phosphatidylcholine complex could not be detected after ultracentrifugation. When added to factor V, cardiolipin (200 mug/ml), produced a rapid 50% decline in activity with a subsequent three-fold increase in the rate of inactivation. No activity was recovered after ultracentrifugation of factor V in the presence of cardiolipin. Saturated phosphatidylethanolamine produced a concentration dependent initial loss of activity, but only a minimal increase in the subsequent rate of inactivation. At 200 mug/ml almost no light complex was detected after ultracentrifugation, but at 800 mug/ml a light complex was observed. This behavior corresponds to the ability of saturated phosphatidylethanolamine to accelerate prothrombin conversion only at very high concentrations. Thus, phospholipids combine with factor V to form complexes which differ in their ability to accelerate prothrombin conversion. The most active species are stable lipoprotein complexes of lower buoyant density than factor V.
D000818:Animals; D002308:Cardiolipins / Q000494:pharmacology; D002417:Cattle; D002499:Centrifugation, Density Gradient; D004355:Drug Stability; D004789:Enzyme Activation; D005165:Factor V / Q000032:analysis* / Q000378:metabolism; D006207:Half-Life; D008823:Micelles; D008968:Molecular Conformation; D008970:Molecular Weight; D010713:Phosphatidylcholines / Q000494:pharmacology; D010714:Phosphatidylethanolamines / Q000494:pharmacology; D010718:Phosphatidylserines / Q000494:pharmacology; D010743:Phospholipids / Q000032:analysis*; D011485:Protein Binding; D011516:Prothrombin / Q000378:metabolism; D013917:Thrombin / Q000096:biosynthesis
D002308:Cardiolipins; D008823:Micelles; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010718:Phosphatidylserines; D010743:Phospholipids; D005165:Factor V; D011516:Prothrombin; D013917:Thrombin