Schoen P, Lindhout T, Willems G, Hemker H C
Department of Biochemistry, University of Limburg, Maastricht, The Netherlands.
Thromb Haemost. 1990 Dec 28;64(4):542-7.
The activation of prothrombin by prothrombinase was investigated in a continuous flow system at 25 degrees C. A glass capillary, containing a continuous phospholipid bilayer attached to the interior surface, was first perfused with factor Va. The factor Va bound to the phospholipid surface functioned as sites for the formation of prothrombinase, when subsequently a factor Xa and prothrombin containing solution was perfused. Under the conditions used, steady-state rates of prothrombin activation were attained after 4 to 15 min. The rates of prothrombinase formation increased with increasing factor Xa concentrations and flow rates, which is compatible with the assembly of prothrombinase being dependent on the flux of factor Xa to the phospholipid-bound factor Va. As long as factor Xa and prothrombin were present in the fluid phase the assembly of prothrombinase was apparently irreversible; during at least 20 min no loss of activity occurred. The steady-state rate of prothrombin activation was dependent on the surface concentration of prothrombinase, at 1.0 microM prothrombin and a shear rate of 82 s-1 the average rate was 870 mol thrombin/min per mol prothrombinase. In contrast to test tube experiments it was observed that in this flow system, the formation of alpha-thrombin is favoured above the formation of meizothrombin (des fragment 1).
在25摄氏度的连续流动系统中研究了凝血酶原酶对凝血酶原的激活作用。首先用因子Va灌注一根玻璃毛细管，该毛细管内表面附着有连续的磷脂双层。当随后灌注含有因子Xa和凝血酶原的溶液时，结合在磷脂表面的因子Va作为凝血酶原酶形成的位点。在所使用的条件下，4至15分钟后达到凝血酶原激活的稳态速率。凝血酶原酶的形成速率随因子Xa浓度和流速的增加而增加，这与凝血酶原酶的组装依赖于因子Xa向结合在磷脂上的因子Va的通量一致。只要因子Xa和凝血酶原存在于液相中，凝血酶原酶的组装显然是不可逆的；至少在20分钟内没有活性损失。凝血酶原激活的稳态速率取决于凝血酶原酶的表面浓度，在凝血酶原浓度为1.0 microM且剪切速率为82 s-1时，平均速率为每摩尔凝血酶原酶每分钟870摩尔凝血酶。与试管实验不同的是，在该流动系统中观察到，α-凝血酶的形成比中间凝血酶（去片段1）的形成更受青睐。
