REGULATORS OF TRANSLATION ELONGATION FACTOR 1 ALPHA

翻译延伸因子1 α（EF-1 α）是一种中心蛋白质， 参与将氨酰-tRNA传递到核糖体， 在蛋白质合成过程中掺入适当的氨基酸。 虽然这种蛋白质是非常丰富的，细胞需要调节 正常细胞生长的EF-1 α活性或水平， EF-α不适当表达导致细胞转化 并发生在许多癌症中。 此外，活动中的变化 EF-1 α的变化发生在衰老过程中，并改变寿命。 这个目标 项目是了解机制，调节效率和 准确的翻译。 有许多不同的机制来确保 mRNA的准确和有效翻译，很明显， 这些机制集中在EF-1 α的作用上。 的核心作用 EF-1 α在延伸中及其与蛋白质和RNA的相互作用， 调节其活性是这一过程中非常重要的因素。 我们 先前的研究支持这一假设， 携带EF-1 α突变的细胞中的保真度是许多因素的结果。 对体内基因表达的不同影响。 当前型号的 伸长率预测这些包括间接影响， 活性EF-1 α x GTP池的变化，从而 核糖体的A位点 然而，EF-1 α也直接影响 这一过程通过将aa-tRNA呈递给核糖体， 同源密码子-反密码子相互作用的确定，以及 动力学校对所需的GTP水解。 的使用 酵母酿酒酵母允许分析翻译 在体内和体外的延长周期。 该提案描述了一个 一套预测或设计用来分析突变的生化研究 EF-1 α对精确基因的直接和间接作用 表情野生型和突变型EF-1 α的纯化和分析 在翻译延伸的部分反应中的蛋白质将 确定体内翻译和生长表型的原因 这些突变。 此外，这些信息还可用于 遗传学方法来识别和表征调节 EF-1 α活性;鸟嘌呤核苷酸交换因子（EF-1 β） 和GTP酶激活因子（核糖体）。 该分析将 还确定了影响或调节尖锐伸长的新因素。 这种协调一致的方法是一种独特的方法， 在许多EF-1 α依赖性步骤中， 高效基因表达

Translation Elongation Factor 1alpha (EF-1alpha) is a central protein involved in delivering aminoacyl-tRNAs to the ribosome and assuring incorporation of the appropriate amino acid during protein synthesis. While this protein is extremely abundant, the cell needs to modulate the activity or levels of EF-1alpha for normal cellular growth since inappropriate expression of EF-alpha results in transformation of cell lines and occurs in many carcinomas. Further, changes in the activity of EF-1alpha occur during aging and alter longevity. The goal of this project is to understand the mechanisms that regulate efficient and accurate translation. There are many different mechanisms to assure the accurate and efficient translation of an mRNA, and it is clear these mechanisms converge on the action of EF-1alpha. The central role of EF-1alpha in elongation and its interactions with proteins and RNAs that modulate its activity are a very important factor in this process. Our previous studies support the hypothesis that changes in translational fidelity in cells harboring mutations in EF-1alpha are a result of many different effects on gene expression in vivo. Current models of elongation predict these include indirect effects that result from changes in the pool of active EF-1alpha x GTP and thus the occupancy of the A-site of the ribosome. However, EF-1alpha also directly affects this process through the presentation of aa-tRNA to the ribosome, the determination of cognate codon-anticodon interactions, and the hydrolysis of GTP required for kinetic proofreading. The use of the yeast Saccharomyces cerevisiae allows analysis of the translation elongation cycle both in vivo and in vitro. This proposal describes a set of biochemical studies of mutations predicted or designed to analyze both direct and indirect effects of EF-1alpha on accurate gene expression. Purification and analysis of wild-type and mutant EF-1alpha proteins in the partial reactions of translation elongation will determine the causes of the in vivo translation and growth phenotypes of these mutations. Furthermore, this information is utilized to apply genetic approaches to identify and characterize factors that modulate EF-1alpha activity; the guanine nucleotide exchange factor (EF-1Beta) and the GTPase activating factor (the ribosome). This analysis will also identify novel factors that affect or regulate acurate elongation. This coordinated approach is a unique method to extend our understanding of the many EF-1alpha dependent steps required for accurate and efficient gene expression.