Structural and Functional Roles of Modules in Protein Architecture

蛋白质结构中模块的结构和功能作用

基本信息

批准号：
02404089
负责人：
GO Mitiko
金额：
$ 24.83万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (A)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1991
项目状态：
已结题

项目摘要

A globular protein consists of compact modules. Module boundaries of proteins are closely correlated with intron positions in the genes encoding the proteins. This fact implies that exon-shuffling in genes is module shuffling in protein level and modules themselves were primitive enzymes. The purpose of this project is to obtain supporting evidences of modules as primitive enzymes and to understand the principle of protein design in nature. Proteins used in this study were barnase that is a bacterial RNase, RNA polymerase II, transcription repressors of procaryotes and ribonuclear proteins in chloroplast, etc. Barnase was decomposed into six modules(M1-M6). Hydrogen bonds were located predominantly within modules in barnase. This fact shows that hydrogen bond formation contributes to determine the compact conformation of modules in barnase architecture. We synthesized modules chemically and carried out experimental study on conformation of each module in solution and its function. Three modules (M2, M3 and M6) were revealed to have an enzymatic function as RNase. This is the first report on the enzymatic function of modules. Our results clearly imply that the modules carried a role as primitive enzymes in early evolution. We also determined the secondary structures of modules M2 and M3 using 2D-NMR and distance geometry. They had the similar secondary structures to those found in intact barnase. Furthermore, we found a barnase-like domain in RNA polymerase II and predicted RNase catalytic function in this region. By other research groups RNase activity of RNA polymerase II was reported. We have suggested that this region might be involved in proof reading, if any. By analysis of module organization in transcription repressors, we found that some of them have two or three helix-turn-helix modules and discussed evolutionary process of repressors through module combinations.

球状蛋白由紧凑的模块组成。蛋白质的模块边界与编码蛋白质的基因中的内含子位置密切相关。这一事实意味着，基因的外显子洗牌是蛋白质水平上的模块改组，模块本身是原始酶。该项目的目的是获得模块作为原始酶的支持证据，并了解自然界的蛋白质设计原理。这项研究中使用的蛋白质是barnase，它是细菌RNase，RNA聚合酶II，procaryotes的转录抑制剂和叶绿体中的核糖核蛋白质等。Barnase分解为六个模块（M1-M6）。氢键主要位于巴尔尼亚酶中的模块中。这一事实表明，氢键的形成有助于确定barnase结构中模块的紧凑构象。我们通过化学合成模块，并就溶液及其功能中每个模块的构象进行了实验研究。揭示了三个模块（M2，M3和M6）具有酶促功能为RNase。这是关于模块酶促功能的第一个报告。我们的结果清楚地表明，模块在早期进化中扮演了原始酶的作用。我们还使用2D-NMR和距离几何形状确定了模块M2和M3的二级结构。它们具有与完整barnase中发现的二级结构相似的。此外，我们在RNA聚合酶II中发现了一个类似巴尔米酶的结构域，并预测了该区域的RNase催化功能。由其他研究组的RNASE活性报告了RNA聚合酶II的RNase活性。我们建议该地区可能参与证明阅读（如果有）。通过对转录阻遏物中模块组织的分析，我们发现其中一些具有两个或三个螺旋 - 螺旋 - 螺旋模块，并通过模块组合讨论了阻遏物的进化过程。