PHOSPHOINOSITIDES IN NORMAL AND ABNORMAL HUMAN RED CELLS

存在于红细胞膜中的磷酸肌醇具有独特的 与许多其他细胞类型相比， 在 红细胞磷酸肌醇起着 与更常见的观察到的信号相反， 转导功能 具体来说，这些脂质已经被 在控制的本地双凹形状的牵连 红细胞以及作为一个组成部分，在骨骼 带4.1和血型糖蛋白之间的复合物。 在这样的背景下， 本提案的重点是了解 磷脂酰肌醇的生理功能和酶学性质 红细胞第一个目标是净化和表征 磷脂酰肌醇4-磷酸激酶。这种酶， 被证明是红细胞骨架的一个组成部分，是一个 控制磷脂酰肌醇水平的关键因素 膜中的4，5-二磷酸。因此，酶将 纯化后，其动力学性质检查上和关闭的 骨架上的这种酶的结合位点将 被识别。 本提案的第二个目标是审查 衰老红细胞的磷酸肌醇代谢。 成熟 红细胞不能合成额外的磷脂酰肌醇， 他们有可能减少这种水平 脂质在他们的生命周期是特别感兴趣的， 老化红细胞的活力。 最近开发的 分离衰老红细胞的程序将是 利用;该程序涉及兔的生物素化 红细胞 衍生化细胞的再输注及其 在体内老化后，通过附着到抗生物素蛋白上而最终恢复 支持.该项目的第三个目标是审查 维持不同的池内的磷酸肌醇 红细胞膜初步研究表明， 部分红细胞磷酸肌醇与 骨骼和这个池子和大部分之间的交换 磷酸肌醇的合成速度很慢，很容易被研究。的 该项目的最终目标是检查未定义的患者， 溶血性贫血的存在异常， 磷酸肌醇途径。

Phosphoinositides present in the erythrocyte membrane have a unique function when compared to many other cell types. In the erythrocyte the phosphoinositides play a structural role in contrast to the more commonly observed signal transduction function. Specifically, these lipids have been implicated in the control of the native biconcave shape of the erythrocyte as well as serving as a component in the skeletal complex between band 4.1 and glycophorin. With this background, the focus of the present proposal is to understand the physiological function and enzymology of the phosphoinositides in erythrocytes. The first goal is to purify and characterize phosphatidylinositol 4-phosphate kinase. This enzyme, which has been shown to be a component of the erythrocyte skeleton, is a critical element in controlling the level of phosphatidylinositol 4,5-bisphosphate in the membrane. Therefore, the enzyme will be purified, its kinetic properties examined both on and off the skeleton, and the binding site for this enzyme on the skeleton will be identified. The second goal of this proposal is to examine the phosphoinositide metabolism of senescent erythrocytes. Mature erythrocytes cannot synthesize additional phosphatidylinositol so that the possibility that they have a decreasing level of this lipid during their life span is of particular interest with regards to the viability of aged erythrocytes. A recently developed procedure for the isolation of senescent erythrocytes will be utilized; this procedure involves the biotinylation of rabbit erythrocytes. The reinfusion of the derivatized cells and their eventual recovery, after in vivo aging, by attachment to an avidin support. The third goal of the project is to examine the maintenance of distinct pools of phosphoinositides within the erythrocyte membrane. Preliminary work has shown that a sizable portion of the erythrocyte phosphoinositides are associated with the skeleton and that the exchange between this pool and the bulk of the phosphoinositides is slow enough to be readily studied. The final goal of the project is to examine patients with undefined hemolytic anemia for the presence of abnormalities in the phosphoinositide pathways.