MEMBRANES AND ACTIVE TRANSPORT OF AMINO ACIDS

易位蛋白、交通ATP酶（或ABC蛋白）、 囊性纤维化跨膜传导调节因子（CFTR），P- 多药耐药（MDR）糖蛋白和细菌周质 渗透酶。 多药耐药是肿瘤的主要问题之一 化疗囊性纤维化是最常见的隐性白种人 疾病 周质渗透酶已被广泛研究， 一个很好的模型系统，用于了解的作用机制， 医学上相关的真核生物超家族成员。 一个这样 通透酶（即组氨酸通透酶）已被详细表征。 如 对于运输ATP酶来说，组氨酸通透酶一般由以下组成： 两个疏水结构域是膜的组成部分，以及两个疏水结构域， 也插入膜中并结合ATP的亲水结构域。 ATP的水解被用作能量来源。 由于CFTR似乎是一个通道，因此确定是否 原核系统也起通道的作用。 这将是一个完全 原核系统的新概念。 从已知的 膜结合复合物，确实有可能疏水结构域 形成一个通道，通过该通道， 膜，ATP水解导致必要的构象 变化 周质渗透酶特有的特征是 存在一个受体，将底物集中在外部 膜结合复合物的表面。 受体向神经元发出信号， 膜结合复合物，导致ATP水解和易位。 在这项研究中将使用的工具是几个重组 系统和几种可测量的酶活性， 功能分析。 作为信道的业务ATP的活动将被 在脂质双层中研究。 信号之间的机制 可溶性受体和膜结合复合物，特别是能量- 偶联组分，将通过使用生物化学反应进行研究 区分不同构象的蛋白质， 有限的蛋白水解和共价标记，并通过遗传分析， 分离出信号传导过程改变的突变体。 类似 生物化学和遗传学程序将被用来研究这种结构 膜结合复合体的一部分 此外， 膜结合复合物将被单独纯化和表征。 二维和三维晶体学都将试图 理解复合体和亚基的结构。 除了解决与作用机制有关的基本问题外， 一般来说，这种原核模型系统的研究将 帮助真核生物研究人员努力解决 与多药耐药性、囊性纤维化、疟疾 寄生虫控制和其他。

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.