(NA+ +K+) - ATPASE--RELATION OF STRUCTURE TO FUNCTION

NaK-ATP酶是质膜的内在酶， 大多数细胞膜表面的正常Na+和K+梯度 是正性肌力作用的受体 洋地黄类药物在心肌中的作用 这一长期目标 项目是确定跨膜离子的分子机制 运动通过这种酶，并阐明机制，控制 其在生理和病理生理条件下的功能。 的 合作建议的重点是功能的重要性， 四级结构的酶，是基于研究结果的 合作研究人员认为，亚基的相互作用， 鸭鼻盐腺酶可能与鸭鼻盐腺酶显着不同 主要研究者广泛使用的酶。 的 建议的比较研究的具体目标是：1.确定 化学计量的活性位点和几个配体结合位点的 鸭盐腺酶，以观察该制剂是否显示半位点 反应性，并将这些化学计量与已知的 肾脏酶的化学计量。2. 为了研究 与鸭子相互作用的几种配体的平衡结合图 盐腺酶，以确定这些是否支持协同结合 配体的酶的底物速度曲线所建议的。3.到 研究哇巴因与鸭盐腺酶的结合，并比较哇巴因与鸭盐腺酶的结合情况。 酶的几种生理配体对哇巴因释放的影响 从盐腺和肾酶，以表征 两种制剂中配体结合位点的多样性。4.到 研究鸭盐腺酶的亚基关联， 化学交联的研究，并比较性能的 膜结合和去污剂溶解的鸭盐腺酶 准备工作5.为了确定这些蛋白亚基的一级序列， 盐腺酶制备亚基的鉴定 参与寡聚化的结构域。

NaK-ATPase is the intrinsic enzyme of the plasma membrane that maintains the normal Na+ and K+ gradients across the surface membranes of most eucaryotic cells, and is the receptor for the positive inotropic actions of digitalis drugs in the myocardium. The long-term objectives of this project are to define the molecular mechanisms for the transmembrane ion movements through this enzyme, and to elucidate the mechanisms that control its function under physiological and pathophysiological conditions. The collaborative proposal is focused on the functional significance of the quaternary structure of the enzyme, and is based on the findings of the collaborating investigator suggesting that the subunit interactions of the duck nasal salt gland enzyme may differ significantly from those of the enzymes that have been used extensively by the Principal Investigator. The specific aims of the proposed comparative studies are: 1. To determine the stoichiometries of the active site and several ligand binding sites of the duck salt gland enzyme to see if this preparation exhibits half-site reactivity, and to compare these stoichiometries with the known stoichiometries of the kidney enzyme. 2. To examine the nature of the equilibrium binding plots for several ligands that interact with the duck salt gland enzyme to determine if these support the cooperative binding of ligands as suggested by the substrate-velocity curves of the enzyme. 3. To study ouabain binding to the duck salt gland enzyme, and to compare the effects of several physiological ligands of the enzyme on ouabain release from the salt gland and the kidney enzymes in order to characterize the multiplicity of the ligand binding sites in the two preparations. 4. To examine the subunit associations of the duck salt gland enzyme through chemical cross-linking studies, and to compare the properties of the membrane-bound and detergent-solubilized duck salt gland enzyme preparations. 5. To determine the primary sequences of the subunits of the salt gland enzyme in preparation for the identification of the subunit domains that are involved in oligomerization.