The Facility for Atomic Mutagenesis

原子诱变设施

基本信息

  • 批准号:
    10063065
  • 负责人:
  • 金额:
    $ 24.79万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2017
  • 资助国家:
    美国
  • 起止时间:
    2017-12-01 至 2022-11-30
  • 项目状态:
    已结题

项目摘要

Ongoing advances in the elucidation of protein structures are leading to the production of large volumes of high-resolution data, in increasingly native environments. However, especially in the case of transmembrane proteins, experimental options for high-resolution functional analyses remain limited, and thus the usefulness of the structures in understanding human neurological diseases likewise remains limited. Moreover, the structures that are obtained represent single, static snapshots of proteins, although these likely have multiple conformations of functional significance. Thus, there is a growing need among the ion channel and membrane biophysics community, which is supported by the NINDS, for reagents that directly report on the functionality and dynamics of membrane proteins in live cells. An elegant solution to these problems is nonsense suppression, a method that makes it possible to encode any type of synthetic amino acid at a site of interest within a protein. These so-called unnatural amino acids can take the form of residues with single atom substitutions, or side-chains with novel fluorescent properties. Although multiple experimental avenues for the encoding of unnatural amino acids exist, each is associated with significant technical challenges. As such, this powerful approach remains inaccessible to most investigators studying molecular neuroscience. In 2014, in response to numerous inquiries by other investigators, the Ahern lab at the University of Iowa (UI) set out to simplify the dissemination of acylated orthogonal tRNAs, key components in nonsense suppression, for experiments involving eukaryotic membrane proteins. To this end, we made improvements to the underlying chemistry, making it more robust and allowing for the encoding of a more chemically diverse set of amino acids. In addition, these new reagents display vastly improved stability profiles thus allowing for easy shipping to laboratories throughout the U.S. With our reagents and guidance, a number of new user groups have successfully applied this previously difficult approach to a variety of membrane proteins relevant to the NINDS mission. Overall, these efforts have produced a high-functioning collaborative outreach service, “The Facility for Atomic Mutagenesis.” This resource will provide broad access to custom reagents for nonsense suppression, and this facility is able to quickly adapt or design synthetic approaches to meet the interests of an application by new users. As the technologies become more standardized and our user group expands, we will scale accordingly, taking advantage of infrastructure present with the UI Carver College of Medicine and local industrial partners such as Integrated DNA Technologies. These collaborations will ultimately support more efficient dissemination of these research tools, to answer diverse questions in molecular neuroscience. Their use will be buoyed by a growing user base, annual training seminars, web-based forums and published protocols in open-access peer-reviewed journals. Our Advisory Board – Kossiakoff, Perozo, Koide, Nakamoto – will ensure efficient stewardship of key resources and the alignment of our strategic vision to NINDS.
在阐明蛋白质结构方面的不断进步导致了大量的 高分辨率数据,在越来越本机的环境中。然而,尤其是在跨膜的情况下 蛋白质,高分辨率功能分析的实验选择仍然有限,因此其有用性 在理解人类神经疾病方面的结构也同样有限。此外, 所获得的结构代表蛋白质的单个静态快照,尽管这些快照可能具有多个 具有功能意义的构象。因此,离子通道和膜之间的需求越来越大 生物物理学社区,由NINDS支持,用于直接报告功能的试剂 以及活细胞中膜蛋白的动态变化。解决这些问题的好办法就是胡说八道 抑制,一种可以在感兴趣的位置编码任何类型的合成氨基酸的方法 在一种蛋白质中。这些所谓的非天然氨基酸可以是单原子残基的形式。 取代,或具有新的荧光性质的侧链。尽管有多个实验途径可以为 存在非天然氨基酸的编码,每一种都伴随着重大的技术挑战。因此,这是 大多数研究分子神经科学的研究人员仍然无法使用强大的方法。2014年,在 在回应其他研究人员的无数询问时,爱荷华大学(UI)的埃亨实验室着手 简化酰化的正交化tRNA的传播,这是无意义抑制的关键成分,用于 涉及真核膜蛋白的实验。为此,我们对底层的 化学,使其更强健,并允许对一组更具化学多样性的氨基进行编码 酸。此外,这些新试剂的稳定性得到了极大的改善,因此便于运输 在我们的试剂和指导下,一些新的用户群体已经 成功地将这一以前困难的方法应用于与NINDS相关的各种膜蛋白 任务。总体而言,这些努力产生了一个高度运作的协作性外联服务--“基金 用于原子诱变。“此资源将提供对定制试剂的广泛访问,以查找无稽之谈 压制,该设施能够快速调整或设计合成方法,以满足 新用户的应用程序。随着技术变得更加标准化,我们的用户群不断扩大,我们将 相应地进行扩展,利用UI Carver医学院和当地的现有基础设施 行业合作伙伴,如集成DNA技术公司。这些协作最终将支持更多 有效地传播这些研究工具,以回答分子神经科学中的各种问题。他们的 用户数量的增长、年度培训研讨会、基于网络的论坛和出版物将提振使用量 开放获取同行评议期刊中的协议。我们的顾问委员会-Kossiakoff,Perozo,Koide,Nakamoto -将确保有效管理关键资源,并确保我们的战略愿景与NINDS保持一致。

项目成果

期刊论文数量(10)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Chemically Acylated tRNAs are Functional in Zebrafish Embryos.
  • DOI:
    10.1021/jacs.2c11452
  • 发表时间:
    2023-01
  • 期刊:
  • 影响因子:
    15
  • 作者:
    Wes Brown;J. Galpin;Carolyn Rosenblum;M. Tsang;C. Ahern;A. Deiters
  • 通讯作者:
    Wes Brown;J. Galpin;Carolyn Rosenblum;M. Tsang;C. Ahern;A. Deiters
Orthogonality of Pyrrolysine tRNA in the Xenopus oocyte.
  • DOI:
    10.1038/s41598-018-23201-z
  • 发表时间:
    2018-03-26
  • 期刊:
  • 影响因子:
    4.6
  • 作者:
    Infield DT;Lueck JD;Galpin JD;Galles GD;Ahern CA
  • 通讯作者:
    Ahern CA
Replacing voltage sensor arginines with citrulline provides mechanistic insight into charge versus shape.
  • DOI:
    10.1085/jgp.201812075
  • 发表时间:
    2018-07-02
  • 期刊:
  • 影响因子:
    0
  • 作者:
    Infield DT;Lee EEL;Galpin JD;Galles GD;Bezanilla F;Ahern CA
  • 通讯作者:
    Ahern CA
Mechanistic insights into robust cardiac I Ks potassium channel activation by aromatic polyunsaturated fatty acid analogues.
芳香族多不饱和脂肪酸类似物强效心脏 IKs 钾通道激活的机制见解。
  • DOI:
    10.1101/2023.01.12.523777
  • 发表时间:
    2023
  • 期刊:
  • 影响因子:
    0
  • 作者:
    Bohannon,BrianaM;Jowais,JessicaJ;Nyberg,Leif;Liin,SaraI;Larsson,HPeter
  • 通讯作者:
    Larsson,HPeter
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Christopher A Ahern其他文献

Christopher A Ahern的其他文献

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{{ truncateString('Christopher A Ahern', 18)}}的其他基金

Chemical biology of voltage-gated cation channels
电压门控阳离子通道的化学生物学
  • 批准号:
    10552311
  • 财政年份:
    2023
  • 资助金额:
    $ 24.79万
  • 项目类别:
A Versatile Chemical-Genetic Approach to Determine Bases for Arrhythmogenesis and Sodium Channelopathies
确定心律失常发生和钠离子通道病基础的多功能化学遗传学方法
  • 批准号:
    10608370
  • 财政年份:
    2022
  • 资助金额:
    $ 24.79万
  • 项目类别:
Restoring Vision with High-Fidelity Nonsense Codon Correction
通过高保真无义密码子校正恢复视力
  • 批准号:
    10550272
  • 财政年份:
    2021
  • 资助金额:
    $ 24.79万
  • 项目类别:
Restoring Vision with High-Fidelity Nonsense Codon Correction
通过高保真无义密码子校正恢复视力
  • 批准号:
    10156779
  • 财政年份:
    2021
  • 资助金额:
    $ 24.79万
  • 项目类别:
Restoring Vision with High-Fidelity Nonsense Codon Correction
通过高保真无义密码子校正恢复视力
  • 批准号:
    10334544
  • 财政年份:
    2021
  • 资助金额:
    $ 24.79万
  • 项目类别:
Restoring Vision with High-Fidelity Nonsense Codon Correction
通过高保真无义密码子校正恢复视力
  • 批准号:
    10407714
  • 财政年份:
    2021
  • 资助金额:
    $ 24.79万
  • 项目类别:
Restoring Vision with High-Fidelity Nonsense Codon Correction
通过高保真无义密码子校正恢复视力
  • 批准号:
    10627046
  • 财政年份:
    2021
  • 资助金额:
    $ 24.79万
  • 项目类别:
Mining the tRNA genome by live-cell imaging
通过活细胞成像挖掘 tRNA 基因组
  • 批准号:
    10005950
  • 财政年份:
    2019
  • 资助金额:
    $ 24.79万
  • 项目类别:
Photochemical determination of sodium channel voltage-dependent gating and composition
钠通道电压依赖性门控和成分的光化学测定
  • 批准号:
    9402276
  • 财政年份:
    2017
  • 资助金额:
    $ 24.79万
  • 项目类别:
Photochemical determination of sodium channel voltage-dependent gating and composition
钠通道电压依赖性门控和成分的光化学测定
  • 批准号:
    10004154
  • 财政年份:
    2017
  • 资助金额:
    $ 24.79万
  • 项目类别:

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