MEMBRANES AND ACTIVE TRANSPORT OF AMINO ACIDS

转运子超家族，交通ATPase(或ABC蛋白)， 囊性纤维化跨膜电导调节因子(CFTR)，P- 多药耐药糖蛋白(MDR)与细菌周质 通行证。多药耐药是癌症的主要问题之一。 化疗和囊性纤维化是最常见的隐性高加索人 疾病。周质透性已被广泛研究并提供了 一个很好的模型系统，可以用来理解 医学上相关的真核超家族成员。这样的一个 渗透酶是组氨酸渗透酶，对其进行了详细的表征。按原样 对于交通ATPase来说，一般来说，组氨酸渗透酶由以下成分组成 作为膜的组成部分的两个疏水结构域，以及两个 也插入到膜中并与ATP结合的亲水结构域。 以三磷酸腺苷的水解物作为能量来源。 由于CFTR似乎是一个通道，因此重要的是确定 原核生物系统也起着通道的作用。这将是一个完全 原核系统的新概念。从已知的结构来看 膜结合的复合体，确实有可能是疏水结构域 的周质渗透层形成一个通道，底物穿过该通道 膜，与三磷酸腺苷的水解导致必要的构象 改变。周质透皮特有的一个特征是 存在将底物集中在外部的受体 膜结合复合体的表面。受体将信号发送到 膜结合复合体，导致三磷酸腺苷的水解和易位。 在这项研究中将使用的工具包括几个重新组合的工具 体外允许的系统和几种可测量的酶活性 功能分析。作为通道的交通ATPase的活动将是 在脂质双分子层中进行研究。之间的信令机制 可溶性受体和膜结合复合体，特别是能量受体- 偶联成分，将利用生化反应进行研究 区分不同构象的蛋白质，例如 有限的蛋白质分解和共价标记，以及通过 分离信号过程发生改变的突变体。类似 生化和遗传程序将被用来研究建筑 与膜结合的复合体。此外，还提供了 膜结合的复合体将被单独提纯和表征。 二维和三维结晶学都将尝试 了解复合体和亚单位的结构。 除了解决与作用机制有关的基本问题外 对于一般的渗透率，对这个原核模型系统的研究将 帮助真核研究人员努力解决 与多药耐药、囊性纤维化、疟疾相关的医疗问题 寄生虫遏制，以及其他。

The superfamily of translocators, traffic ATPases (or ABC proteins), the cystic fibrosis transmembrane conductance regulator (CFTR), the P- glycoprotein of multidrug resistance (MDR), and bacterial periplasmic permeases. Multidrug resistance is one of the major problems in cancer chemotherapy an cystic fibrosis is the most common recessive caucasian disease. Periplasmic permeases have been extensively studied and provide a good model system for understanding the mechanism of action of the medically relevant eukaryotic members of the superfamily. One such permease, the histidine permease, has been characterized in detail. As is true for traffic ATPases in general, the histidine permease is composed of two hydrophobic domains that are integral parts of the membrane, and of two hydrophilic domains that are also inserted into the membrane and bind ATP. Hydrolysis of ATP is used as the energy source. Since CFTR appears to be a channel, it is important to determine whether prokaryotic systems also function as channels. This would be an entirely novel concept for the prokaryotic systems. From the known structure of the membrane-bound complex, it is indeed possible that the hydrophobic domains of periplasmic permeases form a channel through which the substrate crosses the membrane, with ATP hydrolysis resulting in the necessary conformational changes. A characteristic peculiar to periplasmic permeases is the presence of a receptor that concentrates the substrate at the external surface of the membrane-bound complex. The receptor sends a signal to the membrane-bound complex, resulting in ATP hydrolysis and translocation. Among the tools that will be used in this study are several reconstituted systems and several measurable enzymatic activities that permit in vitro assays of function. The activity of traffic ATPases as channels will be investigated in lipid bilayers. The mechanism of signaling between the soluble receptor and the membrane-bound complex, in particular the energy- coupling component, will be studied by the use of biochemical reactions that distinguish between different conformations of proteins, such as limited proteolysis and covalent labeling, and by genetic analysis through the isolation of mutants with altered signaling processes. Similar biochemical and genetic procedures will be used to study the architecture of the membrane-bound complex. In addition, the components of the membrane-bound complex will be purified and characterized individually. Both two- and three-dimensional crystallography will be attempted to understand the structures of both the complex and the subunits. In addition to solving basic questions related to the mechanism of action of permeases in general, the study of this prokaryotic model system will help the efforts of eukaryotic researchers towards a solution of the medical problems related to multidrug resistance, cystic fibrosis, malarial parasite containment, and others.