Using genetic code expansion to investigate the functional dynamics of proteins

利用遗传密码扩展研究蛋白质的功能动力学

• 批准号: 153541321
• 负责人: Professor Dr. Heinz Neumann
• 金额: --
• 依托单位: Fachbereich Chemie- und Biotechnologie
• 依托单位国家: 德国
• 项目类别: Independent Junior Research Groups
• 财政年份: 2009
• 资助国家: 德国
• 起止时间: 2008-12-31 至 2016-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/153541321?language=en
• 关键词: Using; genetic; code; expansion; investigate

Proteins are the key functional parts of the cell. Their dynamic 3D-structure determines their activity. It is controlled by stimuli like the binding of ligands or post-translational modifications. In order to understand how cells function we must elucidate the dynamic behaviour of proteins. Conformational changes in proteins can be detected by introducing biophysical probes, however, this is often technically very difficult. Here, I suggest a strategy to overcome this limitation that introduces unnatural amino acids (UAAs) with unique biophysical properties using components for genetic code expansion (evolved orthogonal ribosomes and tRNA/aminoacyl-tRNA-synthetase pairs), which I have recently created. This will allow to incorporate multiple different UAAs into the same protein and therefore to introduce FRET pairs, which can be used to decipher the dynamics of proteins and its implication for their molecular mechanism of function. I intend to exploit this unique technology to investigate the impact of histone core acetylations, which can only be obtained in pure form by genetically encoding acetyllysine on recombinant histones and have been inaccessible to biochemical analysis so far, on the dynamics of chromatin assembly and remodelling.

蛋白质是细胞的关键功能部分。它们的动态3D结构决定了它们的活动。它受刺激物如配体结合或翻译后修饰的控制。为了了解细胞的功能，我们必须阐明蛋白质的动态行为。蛋白质的构象变化可以通过引入生物物理探针来检测，然而，这在技术上通常非常困难。在这里，我提出了一个策略来克服这一限制，引入非天然氨基酸（UAA）与独特的生物物理特性，使用遗传密码扩展组件（进化的正交核糖体和tRNA/氨酰-tRNA-合成酶对），我最近创建。这将允许将多个不同的UAA掺入同一蛋白质中，从而引入FRET对，其可用于破译蛋白质的动力学及其对它们的分子功能机制的影响。我打算利用这种独特的技术来研究组蛋白核心乙酰化的影响，这只能通过重组组蛋白上的乙酰赖氨酸基因编码以纯的形式获得，迄今为止还无法进行生化分析，对染色质组装和重塑的动力学。

项目成果

Trapping Chromatin Interacting Proteins with Genetically Encoded, UV-Activatable Crosslinkers In Vivo.

用基因编码的、可紫外线激活的交联剂在体内捕获染色质相互作用蛋白

• DOI: 10.1007/978-1-4939-7574-7_16
• 发表时间: 2018
• 期刊: Methods in molecular biology
• 作者: Hoffmann;Neumann;Neumann-Staubitz
• 通讯作者: Neumann-Staubitz

Axial contraction and short-range compaction of chromatin synergistically promote mitotic chromosome condensation

• DOI: 10.7554/elife.10396
• 发表时间: 2015-11-28
• 期刊: ELIFE
• 影响因子: 7.7
• 作者: Kruitwagen, Tom;Denoth-Lippuner, Annina;Barral, Yves
• 通讯作者: Barral, Yves

A Cascade of Histone Modifications Induces Chromatin Condensation in Mitosis

• DOI: 10.1126/science.1244508
• 发表时间: 2014-01-03
• 期刊: SCIENCE
• 影响因子: 56.9
• 作者: Wilkins, Bryan J.;Rall, Nils A.;Neumann, Heinz
• 通讯作者: Neumann, Heinz

Optimized Plasmid Systems for the Incorporation of Multiple Different Unnatural Amino Acids by Evolved Orthogonal Ribosomes

• DOI: 10.1002/cbic.201402033
• 发表时间: 2014-08-18
• 期刊: CHEMBIOCHEM
• 影响因子: 3.2
• 作者: Lammers, Christoph;Hahn, Liljan E.;Neumann, Heinz
• 通讯作者: Neumann, Heinz