MEMBRANE TRANSPORT PROCESSES IN RED BLOOD CELLS

我们的主要目标是深入了解 负责红细胞中电解质的膜运输 以及这些机制在调节细胞体积中的功能。 这项工作的目的是建立关系， Na：K泵复合物的磷酸化，以及 涉及的各种配体，与转运离子的易位。 研究还将致力于确定能源关系， 运输和细胞代谢之间的耦合机制。 我们计划 研究Na与K的泵化学计量比作为细胞Na的函数 尤其是在人体红细胞中。 这些研究报告将 测定了钠、钾的净运动和泵的产电 对通过荧光估计的膜电位的贡献 染色技术 由于细胞体积和代谢（乳酸盐产生） 在许多这些情况下，这些参数也将受到干扰， 随后可能涉及膜 磷酸甘油酸激酶和Na，K-ATP酶定义ATP的膜池。 将一起评价膜池的功能后果 并试图标记包含该池的蛋白质。 我们建议 进一步表征结合发生的32 P-inorg外排 与Na在非偶联Na流出中。 我们还将尝试了解 Na/Na交换和Na-ATP酶活性之间的关系， 在非耦合模式下发生。 此外，我们会进一步研究 ADP +促进的钠泵外排新模式 被ATP抑制的P-inorg，其中P-inorg被转运 与Na。 我们还建议研究改变 膜电位对Na/K的不同部分反应或模式的影响 泵试图识别泵反应中的潜在敏感步骤 周期 要研究的变化包括磷酰基的改变， 电位和Na和K在细胞内的不同梯度 膜的

Our primary aim 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 sidedness 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, especially in human red cells. 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. Functional consequences of the membrane pool will be evaluated together with an attempt to label the proteins comprising the pool. We propose to characterize further the efflux of 32P-inorg that occurs in conjunction with Na in Uncoupled Na efflux. We will also try to understand the relationship between Na/Na exchange and Na-ATPase activities that are known to take place in the Uncoupled mode. In addition, we will study further a New mode of Na efflux carried out by the pump that is promoted by ADP + P-inorg that is inhibited by ATP and in which P-inorg is transported together with Na. We also propose to study the effects of varying the membrane potential on the different partial reactions or modes of the Na/K pump trying to identify potential-sensitive steps in the pump's reaction cycle. Variations to be studied include alterations in the phosphoryl potential and different sets of the gradients of Na and K across the cell's membrane.