MOLECULAR PHYSIOLOGY OF THE SODIUM PUMP

描述（由申请人提供）：本工作的目的是更全面地了解钠泵或Na，K-ATP酶的分子生理学。Na泵将Na和K离子穿过几乎所有哺乳动物细胞的质膜的主动转运与ATP水解的催化偶联。钠泵负责维持正常的液体和电解质平衡，对肾功能和能量重吸收和营养摄取至关重要。在过去的几十年里，我们已经了解了很多关于这种寡聚蛋白如何发挥其重要功能的信息。然而，很少有人知道的控制和调节的过程中，使亚基组装后合成，并作为一个功能单位传递到质膜。在目前的工作中，我们将采用杆状病毒感染的昆虫细胞系统，引入钠泵亚单位到细胞，通常不表达它们。我们将分别和一起研究绵羊α 1-亚基（1000个氨基酸的多位膜蛋白）和绵羊β 1-亚基（约300个氨基酸的蛋白质和单一膜交叉）的表达，并使用细胞分级分离和代谢标记策略研究控制其在内质网中组装并通过高尔基体传递到质膜的因素。通过利用一系列的突变，在每个亚单位，我们将调查的分子决定因素的作用，影响传递到质膜，其在膜中的稳定性及其与细胞骨架元素的相互作用。钠泵是一种重要的转运系统，也是洋地黄的作用部位，洋地黄是心功能不全最广泛使用的治疗方法。当我们更好地了解细胞膜中Na泵活性的调节时，将大大有助于改善心脏功能和改善肾功能受损的治疗前景。

DESCRIPTION (provided by applicant): The objective of the present work is to obtain a more complete understanding of the molecular physiology of the Na pump or Na,K-ATPase. The Na pump couples the active transport of Na and K ions across the plasma membrane of almost all mammalian cells to the catalysis of ATP hydrolysis. The Na pump is responsible for maintaining normal fluid and electrolyte balance and is essential for renal function and energising reabsorbtion and nutrient uptake. During the past several decades we have learned much about how this oligomeric protein carries out its important function. However, little is known about the control and regulation of the processes which enable the subunits to assemble after synthesis and be delivered as a functioning unit to the plasma membrane. In the present work we will employ the baculovirus-infected insect cell system to introduce Na pump subunits into cells which do not normally express them. We will examine the expression of the sheep alpha1 -subunit (a polytopic membrane protein of 1000 amino acids) and sheep Beta1-subunit (a protein of about 300 amino acids and a single membrane crossing) separately and together and using cell fractionation and metabolic labelling strategies investigate the factors which control their assembly in the endoplasmic reticulum and delivery to the plasma membrane via the golgi apparatus. By utilizing a series of mutations in each sub-unit we will investigate the role of molecular determinants which affect delivery to the plasma membrane, its stability in the membrane and its interactions with cytoskeletal elements. The Na pump is an essential transport system and the site of action of digitalis, the most widely used therapy in cardiac insufficiency. Prospects for improved therapies for cardiac function and for improving impaired renal function will be greatly aided when we have a better understanding of the regulation of the activity of the Na pump in cell membranes.