MOLECULAR BIOLOGY AND BIOCHEMISTRY OF PHOTOSYSTEM II

该项目的长期目标是使用分子工具， 生物学和生物化学，以了解各种基因及其 调节色素-蛋白质复合物功能的产物， 光系统EL一个更直接的目标是了解功能性的 许多非常小的蛋白质（< 10 kDa）在这种多蛋白中的作用 我们的实验生物体是一种单细胞的， 蓝藻，集胞藻6803。 在过去，我们使用“反向 遗传学的方法来创建特定的靶向突变， 编码集胞藻6803的各种PSII蛋白的基因。 我们现在 计划使用类似的方法在基因中创建定点突变体 编码这种蛋白质复合物的小蛋白质。 由于与反应中心的结构和功能相似， 第一种膜结合多蛋白复合物，其X射线晶体数据具有 变得可用。 然而，反应中心复合物不包含 任何小的蛋白质成分。有趣的是，这些小蛋白质 许多膜蛋白复合物的普遍存在的组分，例如，细胞色素 氧化酶。 这些小蛋白的结构和/或功能作用 然而，并没有被清楚地理解。e本项目中的拟议研究 因此，将扩大我们对这种小的功能的一般理解， 所有类似蛋白质复合物中的蛋白质。 这些研究还将 改进今后工作中可能采用的技术和概念， 膜蛋白复合物在生物医学上很重要。 生物化学、遗传学和重组DNA技术的结合 将被用来研究不同突变的影响，在该网站定向 在这段时间里创造的变种人 此外，我们将 分离随机诱变PSII缺陷突变体。 特别 感兴趣的将是使用转座子元件Tn 5 tac 1来创建 条件（IPTG，乳糖类似物，-依赖性）突变体， PSII活动。 使用这种通用的遗传工具和简单的 集胞藻6803的转化系统将帮助我们识别新的 结构基因座，更重要的是，调节基因座， PSII复合物的生物起源、稳定性以及功能。

The long term objective of this project is to use tools of molecular biology and biochemistry to understand the roles of various genes and their products in the regulation of function of a pigment - protein complex, Photosystem EL A more immediate objective is to understand the functional role of a number of very small proteins (< 10 kDa) in this multiprotein complex Our experimental organism is a unicellular, transformable cyanobacterium, Synechocystis 6803. In the past, we have used 'reverse genetics' approaches to create specific targeted mutations in a number of genes that encode various PSII proteins of Synechocystis 6803. We now plan to use similar approaches to create site directed mutants in genes encoding small proteins of this protein complex The PSII complex has structural as well as functional similarities with a reaction center, the first membrane bound multiprotein complex whose Xray crystal data have become available. However, the reaction center complex does not contain any small protein component. Interestingly, such small proteins are ubiquitous components of many membrane protein complexes, e.g., cytochrome oxidase. The structural and/or functional roles of such small proteins are, however, not clearly understood. e proposed studies in this project will thus, broaden our general understanding of the function of such small proteins in all protein-complexes of similar kind. These studies will also refine techniques and concepts that may be employed in future work on membrane protein complexes that are biomedically important. A combination of biochemical, genetic and recombinant DNA techniques will be used to study the effects various mutations in the site directed mutants created during this proposal period. In addition, we will to isolate randomly mutagenized PSII deficient mutants. Of particular interest will be the use of a transposon element, Tn5tac 1, to create conditional (IPTG, a lactose analog, -dependent) mutants that impaired in their PSII activities. Use of such versatile genetic tools and the facile transformation system Synechocystis 6803 will help us in identifying novel structural and, more important, regulatory loci that influence the biogenesis, stability, as well as function, of the PSII complex.