Structure and Function of the Plant Vacuolar H+-ATPase

本项目的目的是表征植物液泡H+- atp酶的结构和功能。该项目的具体目标是：(i)通过进一步纯化和免疫沉淀确定完整的亚基组成；（ii）通过分离和识别外围亚基来研究亚基组织；（iii）通过克隆和测序编码16kda蛋白脂的基因，推断初级结构，然后确定选定亚基的功能域；（iv）通过重组和分子遗传学方法检测16 kDa蛋白脂的功能；(v)定位atp酶的亚细胞分布；（vi）开始对选定的atp酶亚基的生物合成进行初步研究。植物细胞含有一个大的中央液泡，占细胞体积的80%。它们对维持细胞膨胀和离子和代谢物的储存很重要。通过液泡膜（液泡体）的运输过程由电化学梯度控制，电化学梯度部分由液泡体局部液泡atp酶（v - atp酶）决定，这是一种质子泵送atp酶（H+- atp酶）。v - atp酶代表了存在于所有真核细胞的膜系统中的一类atp酶。因此，对植物液泡atp酶的结构和功能的研究与所有高等生物的膜系统功能有关。

The purpose of this project is to characterize the structure and function of the plant vacuolar H+-ATPase. The specific goals of the project are (i) to determine the complete subunit composition by further purification and immunoprecipitation; (ii) to study the subunit organization by dissociating and identifying the peripheral subunits; (iii) to deduce the primary structure and later to define the functional domains of selected subunits by cloning and sequencing the gene encoding the 16 kDa proteolipid; (iv) to test the function of the 16 kDa proteolipid by reconstitution and molecular genetic methods; (v) to localize the subcellular distribution of the ATPase; and (vi) to begin preliminary studies on the biosynthesis of selected ATPase subunits. Plant cells contain a large central vacuole that occupies up to 80% of the cell volume. They are important for maintaining cell turgor and for the storage of ions and metabolites. Transport processes across the vacuolar membrane (tonoplast) are controlled by electrochemical gradients that are determined in part by the tonoplast localized vacuolar ATPase (V-ATPases) which is a proton-pumping ATPase (H+-ATPase). The V-ATPases represent a class of ATPase that is present in the endomembrane system of all eukaryotic cells. Thus, investigations on the structure and function of the plant vacuolar ATPase has relevance to the function of the endomembrane system in all higher organisms.