Kinetics and thermodynamics of annexin A1 binding to solid-supported membranes: A QCM study
Kinetics and thermodynamics of annexin A1 binding to solid-supported membranes: A QCM study
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DOI：
10.1021/bi025951z
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发表时间：
2002-08-06
期刊：
影响因子：
2.9
通讯作者：
Steinem, C
中科院分区：
文献类型：
--
作者：
Kastl, K;Ross, M;Steinem, C
利用石英晶体微天平(QCM)技术，在5 MHz石英板上的金电极上固定了两层膜，定量了膜联蛋白Al与脂膜的相互作用。固体支持的脂质双层由化学吸附在金表面的第一层辛硫醇和通过囊泡融合获得的物理吸附的磷脂单分子层组成。这个实验装置使我们能够第一次确定膜联蛋白Al与含磷脂酰丝氨酸的层结合的速率常数和亲和力常数作为溶液中钙离子浓度和固体支撑双层外层中胆固醇含量的函数。结果表明，当Ca~(2+)浓度从1 mM降至100微米时，膜联蛋白A与膜的结合率显著增加，与胆固醇含量无关。然而，膜中胆固醇的存在极大地改变了亲和常数。在不含胆固醇的1-paimitoyl-2-oleoyl-sn-glycero-3-phosphocholine(POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine(POPS)膜中，结合常数随钙离子浓度的降低而降低，但在有胆固醇存在时，结合常数仍然很高。
By means of the quartz crystal microbalance (QCM) technique, the interaction of annexin Al with lipid membranes was quantified using solid-supported bilayers immobilized on gold electrodes deposited on 5 MHz quartz plates. Solid-supported lipid bilayers were composed of a first octanethiol monolayer chemisorbed on gold and a physisorbed phospholipid monolayer obtained from vesicle fusion. This experimental setup enabled us to determine for the first time rate constants and affinity constants of annexin Al binding to phosphatidylserine-containing layers as a function of the calcium ion concentration in solution and the cholesterol content within the outer leaflet of the solid-supported bilayer. The results reveal that a decrease in Ca2+ concentration from 1 mM to 100 muM significantly increases the rate of annexin A] binding to the membrane independent of the cholesterol content. However, the presence of cholesterol in the membrane altered the affinity constants considerably. While the association constant decreases with decreasing Ca2+ concentration in the case of 1-paimitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine (POPS) membranes lacking cholesterol, it remains high in the presence of cholesterol.