GENETIC APPROACHES TO PROTEIN-NUCLEIC ACID INTERACTIONS

蛋白质-核酸相互作用的遗传学方法

• 批准号: 2174119
• 负责人: PAUL R SCHIMMEL
• 金额: $ 27.71万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 1977
• 资助国家: 美国
• 起止时间: 1977-03-01 至 1997-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2174119
• 关键词: Escherichia coli; Saccharomyces cerevisiae; X ray crystallography; acylation; alanine; aminoacid tRNA ligase; aminoacyl tRNA; biochemical evolution; chemical fingerprinting; crosslink; enzyme induction /repression; enzyme mechanism; enzyme structure; genetic manipulation; glutamine; hybrid enzyme; isoleucine; methionine; mutant; nucleic acid sequence; protein engineering; site directed mutagenesis; temperature sensitive mutant; transfer RNA

Aminoacyl transfer RNA synthetases arose early in evolution and established the rules of the genetic code through their specific aminoacylations of transfer RNAs. the enzymes are organized into two classes of ten enzymes each. These two classes appear to have no historical relationship to each other. Experiments are proposed to investigate and manipulate the tRNA specificity of class I and class II enzymes. These synthetases include E. coli alanine, glutamine, isoleucine and methionine tRNA synthetases, and S. cerevisiae glutamine tRNA synthetase. Recent experiments have shown that the amino acid acceptor helix is a major determinant of the identity of at least some tRNAs. Functional synthetase-tRNA acceptor helix contacts will be identified and manipulated through analysis of mutant enzymes which are selected for specific alterations in their acceptor helix interactions. A minimalist aminoacyl tRNA synthetase will also be designed and subjected to mutagenesis and selection for functional activity. Additional efforts will be directed at manipulations of tRNA recognition specificity by introducing the fewest possible amino acid or nucleotide substitutions, in some cases taking advantage of recent x-ray crystallographic structural information.

氨酰转移RNA合成酶在进化早期出现， 建立了遗传密码的规则， 转移RNA的氨酰化。 酶被组织成两种 每类十种酶。 这两个类似乎没有 彼此的历史关系。 实验被提议为 研究和操纵I类和II类的tRNA特异性， II酶。 这些合成酶包括E.大肠杆菌丙氨酸，谷氨酰胺， 异亮氨酸和甲硫氨酸tRNA合成酶，和S.酿酒谷氨酰胺 tRNA合成酶。 最近的实验表明， 受体螺旋是至少一些人的身份的主要决定因素， tRNA。 功能性合成酶-tRNA受体螺旋接触将是 通过分析突变酶来鉴定和操作， 选择它们的受体螺旋相互作用中的特定改变。 还将设计最低限度的氨酰tRNA合成酶， 进行功能活性的诱变和选择。 另外的努力将针对tRNA识别的操纵 通过引入尽可能少的氨基酸或核苷酸来实现特异性 替代，在某些情况下利用最近的X射线 晶体结构信息