Expanding the Genetic Code with Phosphothreonine and Phosphotyrosine

用磷酸苏氨酸和磷酸酪氨酸扩展遗传密码

• 批准号: 0950474
• 负责人: Dieter Soll
• 金额: $ 54.94万
• 依托单位: Yale University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-15 至 2014-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0950474&HistoricalAwards=false
• 关键词: Expanding; Genetic; Code; Phosphothreonine; Phosphotyrosine

Synthesis of proteins containing amino acids outside of the genetic code has recently resurfaced as a topic of much interest and presents a great challenge intellectually and technically. Reprogramming the genetic code requires two key events: (i) The creation of a new efficient 'orthogonal' aminoacyl tRNA synthetase:tRNA pair able to generate solely a non-canonical aminoacyl tRNA. (ii) The ability to specify the position in which this amino acid will be inserted in the protein by recoding a particular mRNA codon. Based on the natural existence of two archaeal aminoacyl-tRNA synthetase-like proteins (O-phosphoseryl tRNA synthetase with its cognate tRNACys and pyrrolysyl-tRNA synthetase with its cognate tRNAPyl) research will be initiated to develop an Escherichia coli system for the in vivo co translational insertion of phosphothreonine and phosphotyrosine at pre-determined positions in a polypeptide based on the nonsense codons UGA and UAG in a messenger RNA. Given the importance of phosphothreonine and phosphotyrosine in eukaryotic proteins essential for cell proliferation, cellular signaling, and cell death, a robust method of making proteins containing phospho-amino acids will have wide application and great significance. The broader impacts resulting from this project are: (i) The experimental tools to be developed will be widely employed in the future by other researchers in the life sciences. (ii) Development of a viable patent will contribute to commercial technology. (iii) This project will provide interdisciplinary training for undergraduate and postdoctoral students.

含有遗传密码之外的氨基酸的蛋白质的合成最近重新出现，作为一个非常感兴趣的话题，并提出了一个巨大的挑战，智力和技术。重新编程遗传密码需要两个关键事件：（i）创建一个新的有效的“正交”氨酰tRNA合成酶：tRNA对，能够单独产生非规范的氨酰tRNA。(ii)通过重新编码特定的mRNA密码子来指定该氨基酸插入蛋白质的位置的能力。基于两种古细菌氨酰-tRNA合成酶样蛋白（O-磷酸丝氨酰tRNA合成酶及其同源tRNACys和吡咯赖氨酰-tRNA合成酶及其同源tRNAPyl）的天然存在，将启动研究以开发用于基于信使RNA中的无义密码子UGA和UAG在多肽中的预定位置处进行磷酸苏氨酸和磷酸酪氨酸的体内共翻译插入的大肠杆菌系统。鉴于磷酸苏氨酸和磷酸酪氨酸在细胞增殖、细胞信号传导和细胞死亡所必需的真核蛋白质中的重要性，制备含有磷酸氨基酸的蛋白质的稳健方法将具有广泛的应用和重大意义。这一项目产生的更广泛影响是：（一）将要开发的实验工具将在今后被生命科学领域的其他研究人员广泛使用。(ii)开发可行的专利将有助于商业技术。(iii)该项目将为本科生和博士后学生提供跨学科培训。