ANALYSIS OF CARDIAC GLYCOSIDE AFFINITY IN CULTURED CELLS

Na，K-泵(即Na，K-ATPase)被认为是主要的 洋地黄的药理受体，如哇巴因。这是一个 从细胞中排出钠离子的固有膜蛋白复合体 并以代谢为代价从外部吸收K+ 能量。它的周转和由此产生的电化学梯度 是造成整个等离子体的电势差异的原因 膜和间接调节离子平衡，细胞体积，和 盐分和水的上皮性运输。Na，K泵是 被心脏糖苷抑制，并且有变异的 哇巴因在组织之间的亲和力可能高达500倍。 这种可变性的基础还没有被很好地理解，这 该项目将检验哇巴因变异的假设 Na，K泵的亲和力是由细胞特异性引起的 其组成亚基的不同亚型的表达。这个 亚基组成和所表达的哇巴因亲和力将为 用RNA杂交法检测多个培养细胞系 分析和酶分析来关联特异性的存在 具有给定结合亲和力的异构体。DNA介导的基因转移 将用于将异构体导入受体细胞系 直接观察它们对哇巴因亲和力的影响。从这些 哇巴因组织变异的分子基础研究 亲和力将被阐明。 为了实施这个项目，候选人将利用 广泛的背景，包括比较生物化学， 运输生理学和分子生物学。RCDA将确保 候选人可以继续这种多学科的方法。 进行这项工作的部门是 在运输生理和运输领域获得国际认可 提供了一个理想的环境来开发独立的 研究生涯。它位于一个主要的健康科学中心内 应进一步刺激创新研究的发展 Na，K泵的功能和调节。

The Na, K-pump (ie., Na, K-ATPase) is thought to be the major pharmacological receptor for digitalis such as ouabain. It is an intrinsic membrane protein complex that extrudes Na+ from the cell and absorbs K+ from the exterior at the expense of metabolic energy. Its turnover and the resulting electrochemical gradients are responsible for the potential difference across the plasma membrane and indirectly mediate ionic balance, cellular volume, and epithelial transport of salts and water. The Na, K-pump is inhibited by cardiac glycosides, and there is variation in the ouabain affinity between tissues that may be as great as 500-fold. The basis for this variability is not well understood, This project will test the hypothesis that the variation in ouabain affinity of the Na, K-pump is caused by the cell-specific expression of different isoforms of its constituent subunits. The subunit composition and the expressed ouabain affinity will be examined in a number of cultured cell lines by RNA hybridization analysis and enzymatic assays to correlate the presence of specific isoforms with a given binding affinity. DNA-mediated gene transfer will be used to introduce the isoforms into recipient cell lines to observe directly their effects on ouabain affinity. From these studies, the molecular basis of the tissue variability in ouabain affinity will be elucidated. To carry out this project, the candidate will take advantage of a broad background that has included comparative biochemistry, transport physiology, and molecular biology. An RCDA will ensure that the candidate can continue this multidisciplinary approach. The department in which the work will be conducted is internationally recognized in the field of transport physiology and provides an ideal environment in which to develop an independent research career. Its location within a major health science center should further stimulate the development of innovative research in the function and regulation of the Na, K-pump.