School of Biological Sciences, Queen's University Belfast, Medical Biology Centre, Belfast, UK.
Mol Cell Biochem. 2011 Feb;348(1-2):109-15. doi: 10.1007/s11010-010-0644-x. Epub 2010 Nov 16.
Prothrombin interacts with phosphatidylserine containing platelet membranes via its N-terminal, γ-carboxyglutamate (gla) residue-rich domain. Once bound it is cleaved to form the active protease, thrombin (factor IIa). Human prothrombin was cleaved with cathepsin G in the absence of calcium and magnesium ions. Under these conditions, the gla domain was removed. Phospholipid protected the protein from this proteolytic event, and this suggests that a conformational change may be induced by interaction with phospholipids. Binding of prothrombin to a surface containing 20% phosphatidylserine/80% phosphatidylcholine was detected by surface plasmon resonance, whereas no interaction with gla-domainless prothrombin was observed. Binding of intact prothrombin in the presence of calcium ions showed complex association kinetics, suggesting multiple modes of initial interaction with the surface. The kinetics of the dissociation phase could be fitted to a two-phase, exponential decay. This implies that there are at least two forms of the protein on the surface one of which dissociates tenfold more slowly than the other. Taken together, these data suggest that, on binding to a membrane surface, prothrombin undergoes a conformational change to a form which binds more tightly to the membrane.
凝血酶原通过其 N 端富含 γ-羧基谷氨酸（gla）残基的结构域与含有磷脂酰丝氨酸的血小板膜相互作用。一旦结合，它就会被切割形成活性蛋白酶，即凝血酶（因子 IIa）。在没有钙离子和镁离子的情况下，用人组织蛋白酶 G 切割人凝血酶原。在这些条件下，gla 结构域被去除。磷脂可以保护蛋白质免受这种蛋白水解事件的影响，这表明与磷脂的相互作用可能会诱导构象变化。通过表面等离子体共振检测到凝血酶原与含有 20%磷脂酰丝氨酸/80%磷脂酰胆碱的表面的结合，而没有观察到与缺乏 gla 结构域的凝血酶原的相互作用。在钙离子存在下，完整凝血酶原的结合表现出复杂的关联动力学，表明与表面的初始相互作用有多种模式。解吸相的动力学可以拟合为两相指数衰减。这意味着在表面上至少有两种形式的蛋白质，其中一种的解离速度比另一种慢十倍。综上所述，这些数据表明，凝血酶原在与膜表面结合时，会发生构象变化，形成与膜结合更紧密的形式。
