Walker F J
J Biol Chem. 1986 Aug 15;261(23):10941-4.
The apparent molecular weight of functional protein S in citrated plasma was observed to be between 115,000 and 130,000 as measured by sedimentation equilibrium in the air-driven ultracentrifuge. The molecular weight of the functional protein decreased to approximately 62,000 when copper ions were added to the plasma. This suggested the presence of a protein S-binding protein in plasma, which was confirmed by gel filtration experiments. Frontal analysis of plasma indicated that functional protein S could exist in as many as three forms. Addition of copper ions to plasma reduced the number of forms to one. In order to isolate the binding protein, plasma was fractionated first on a column of immobilized iminodiacetic acid that had been equilibrated with copper ions. The proteins that eluted in a 0.6 M NaCl wash were passed over a column of protein S immobilized on agarose beads. A protein, eluted in the 0.6 M NaCl wash, was observed to bind to protein S in gel filtration experiments. When added to plasma depleted of both protein S and the binding protein, the binding protein was observed to enhance the anticoagulant activity of activated protein C only in the presence of protein S. Protein S-binding protein was also observed to enhance the rate of factor Va inactivation by activated protein C and protein S.
通过空气驱动超速离心机中的沉降平衡法测定，枸橼酸盐血浆中功能性蛋白S的表观分子量在115,000至130,000之间。当向血浆中加入铜离子时，功能性蛋白的分子量降至约62,000。这表明血浆中存在一种蛋白S结合蛋白，凝胶过滤实验证实了这一点。血浆的前沿分析表明，功能性蛋白S可能以多达三种形式存在。向血浆中加入铜离子可将形式数量减少至一种。为了分离结合蛋白，首先将血浆在已用铜离子平衡的固定化亚氨基二乙酸柱上进行分级分离。在0.6M NaCl洗脱液中洗脱的蛋白质通过固定在琼脂糖珠上的蛋白S柱。在凝胶过滤实验中观察到，在0.6M NaCl洗脱液中洗脱的一种蛋白质与蛋白S结合。当将其添加到既缺乏蛋白S又缺乏结合蛋白的血浆中时，仅在存在蛋白S的情况下观察到结合蛋白增强活化蛋白C的抗凝活性。还观察到蛋白S结合蛋白增强活化蛋白C和蛋白S使因子Va失活的速率。
