MEMBRANE TRANSPORT PROCESSES IN RED BLOOD CELLS

本建议中概述的工作的主要目的是深入了解 进入负责膜运输的分子机制， 红细胞中的电解质以及这些机制在 调节细胞体积。 这项工作旨在建立 Na：K泵复合物的磷酸化与 参与的各种配体的作用侧， 运输的离子。 研究还将致力于定义 运输和细胞之间的能量关系和耦合机制 新陈代谢. 我们计划研究Na与K的泵化学计量， 细胞钠浓度的函数，以及磷酰基的变化 潜力 这些研究将进行测量净钠和钾 运动和泵对膜的生电作用 通过荧光染料技术估计的电势。 由于细胞 体积和代谢（乳酸盐产生）在许多情况下受到干扰， 在这些情况下，这些参数也将与 可能涉及膜磷酸甘油酸激酶和Na，K-ATP酶 形成ATP的膜池。 我们还计划研究 K及其替代物、强心苷和核苷的特性 完整的膜和膜组件，以努力定义更多 正是这种结构和分子事件 膜转运 此外，我们会继续进行有关的研究 通过识别与Ca诱导的K 通过监测重封血影中Ca水平来研究人红细胞的转运 与包埋砷剂III，膜电位和钾渗透性; 此外，我们计划标记膜位点以确定它们的数量， 可能有助于它们的分离和随后的表征。

The primary aim of the work outlined in this proposal is to gain insight into the molecular mechanisms responsible for the membrane transport of electrolytes in red blood cells and the function these mechanisms have in the regulation of cell volume. The work is oriented toward establishing the relationship between phosphorylation of the Na:K pump complex, and the sideness of action of various ligands involved, with the translocation of the transported ions. Studies will also be directed toward defining the energy relationships and coupling mechanisms between transport and cell metabolism. We plan to study the pump stoichiometry of Na to K as a function of cell Na concentration, and changes in the phosphoryl potential. These studies will be carried out measuring net Na and K movements and the pump's electrogenic contribution to the membrane potential estimated by means of a fluorescence dye technique. Since cell volume and metabolism (lactate production) are perturbed under many of these situations, these parameters will also be followed together with the possible involvement of membrane phosphoglycerate kinase and Na,K-ATPase defining a membrane pool of ATP. We also plan to study the binding characteristics of K and its surrogates, cardiac glycosides and nuclotides to intact membranes and membrane components in an effort to define more precisely the substructure and the molecular events associated with membrane transport. In addition, we will continue our studies concerned with identifying the membrane process associated with Ca-induced K transport of human red cells by monitoring in resealed ghosts the Ca levels with entrapped arsenaoz III, membrane potentials and K permeability; in addition, we plan to label the membrane sites to determine their number and possibly to help in their isolation and subsequent characterization.