GENETICS OF FLAVIN DEHYDROGENASE-MEMBRANE INTERACTIONS

本研究的目的是确定黄素分解酶 与沙门氏菌中的膜结合电子传递链相互作用 鼠伤寒。 这项工作将侧重于两个主要专题： 1. put操纵子的调控。 看跌操纵子具有独特的控制作用 这一机制似乎是由putA基因产物脯氨酸介导的 氧化酶。 当外源脯氨酸可用时，脯氨酸氧化酶结合到 膜在那里它的功能作为黄素脱氢酶;但在缺乏 脯氨酸，它积累在细胞质中，在那里它作为一个自体 阻遏物 确定脯氨酸氧化酶如何控制其自身的表达， 帮助我们了解其他膜结合调节蛋白的工作原理。 在 此外，putA突变的调节特性提供了一种遗传学机制， 选择膜相互作用突变体。 2.黄素氧化酶-膜相互作用。 黄素氧化酶必须 与特定的膜位点结合，以便与 电子传递链。 但是，不知道是什么组件 它们与电子传递链的直接相互作用， 它们与细胞膜相互作用。 遗传学研究的相互作用 具有电子传递链的黄素氧化酶将提供一种 了解蛋白质如何与细胞膜相互作用的新方法 vivo. 所有涉及的组件和特定的相互作用 可以通过隔离和表征两个独特的类来定义所需的 脱氢酶突变：阻止脱氢酶 与膜相互作用，但不消除酶的活性， 脱氢酶在体外，和突变，改变与脱氢酶， 它仍然可以结合膜，但不能正确地与膜相互作用， 电子传递链 这些基因和生化分析 突变体将指示哪些膜组分直接与 以及它们之间需要何种相互作用 黄素化酶和膜电子传递链。 等 突变体应该提供洞察一般的生物学问题， 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.