GENETICS OF FLAVIN DEHYDROGENASE-MEMBRANE INTERACTIONS

这项研究的目的是确定黄素脱氢酶如何 沙门氏菌与膜结合电子传递链的相互作用 鼠伤寒杆菌。这项工作将主要围绕两个主题展开： 1.对看跌期权操纵者的监管。PUT操纵子有一个独特的控制 这一机制似乎是由PuTA基因产物--脯氨酸介导的 氧化物酶。当外源脯氨酸可用时，脯氨酸氧化酶结合到 作为黄素脱氢酶功能的膜；但在缺乏 脯氨酸，它在细胞质中积累，在那里它扮演着自体 抑制者。确定脯氨酸氧化酶如何控制自己的表达可能 帮助我们了解其他膜结合的调节蛋白是如何工作的。在……里面 此外，PUTA突变的调节特性提供了一种基因 膜相互作用突变体的筛选。 2.黄素脱氢酶-膜相互作用。黄素脱氢酶必须 与特定的膜部位结合以便与之正确地相互作用 电子传输链。不过，目前还不知道是什么成分(S) 它们直接或如何相互作用的电子传输链 它们与膜相互作用。相互作用的遗传学研究 黄素脱氢酶与电子传递链将提供 了解蛋白质如何与膜相互作用的新方法 活着。所有涉及的组件和特定的交互 Required可以通过分离和表征两个唯一的类来定义 脱氢酶突变：阻止脱氢酶 与膜相互作用，但不消除酶活性 脱氢酶在体外，以及随着脱氢酶而改变的突变 它仍然可以与膜结合，但不能与 电子传输链。这些细菌的遗传和生化分析 突变体将指示哪些膜组件(S)直接与其相互作用 黄素脱氢酶与黄素脱氢酶的相互作用 黄素脱氢酶和膜电子传递链。是这样的 突变体应该提供对一般生物学问题的洞察，即如何 外周膜蛋白与膜相互作用。

The objective of this study is to determine how flavin dehydrogenases interact with the membrane-bound electron transport chain in Salmonella typhimurium. This work will focus on two main topics: 1. Regulation of the put operon. The put operon has a unique control mechanism that seems to be mediated by the putA gene product, proline oxidase. When exogenous proline is available, proline oxidase binds to the membrane where it functions as flavin dehydrogenase; but in the absence of proline, it accumulates in the cytoplasm where it acts as an autogenous repressor. Determining how proline oxidase controls its own expression may help us understand how other membrane-bound regulatory proteins work. In addition, the regulatory properties of putA mutations provide a genetic selection for membrane interaction mutants. 2. Flavin dehydrogenase-membrane interactions. Flavin dehydrogenases must associate with specific membrane sites in order to interact properly with the electron transport chain. However, it is not known what component(s) of the electron transport chain they interact with directly or even how they interact with the membrane. Genetic studies on the interaction of flavin dehydrogenases with the electron transport chain will provide a novel approach for understanding how proteins interact with membranes in vivo. All of the components involved and the specific interactions required can be defined by isolating and characterizing two unique classes of dehydrogenase mutations: mutations that prevent the dehydrogenase from interacting with the membrane but don't eliminate enzymatic activity of the dehydrogenase in vitro, and mutations that alter with dehydrogenase so that it can still bind the membrane but can't properly interact with the electron transport chain. Genetic and biochemical analysis of these mutants will indicate what membrane component(s) directly interact with flavin dehydrogenases and what kinds of interactions are required between flavin dehydrogenases and the membrane electron transport chain. Such mutants should provide insight into the general biological problem of how peripheral membrane proteins interact with the membrane.