RUI: Development of a Biocompatible Visible-Light-Catalyzed Thiol-Ene Coupling Reaction to Study Redox Active Disulfide Bonds.

RUI：开发生物相容性可见光催化硫醇-烯偶联反应来研究氧化还原活性二硫键。

• 批准号: 1807218
• 负责人: Andrew Wommack
• 金额: $ 17.25万
• 依托单位: High Point University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1807218&HistoricalAwards=false
• 关键词: RUI; Development; Biocompatible; Visible; Light

The Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Andrew J. Wommack from High Point University to examine the functional and structural consequences of sulfur-sulfur or disulfide bonds in proteins. The disulfide bond serves important roles in biology ranging from structural effects in proteins to cell signaling and control of metabolic pathways. This sulfur-sulfur chemical bond can be found in opened or closed states under different biological conditions, such as during different physiological stresses or within unique cellular locations. To study the functional impact of the disulfide bond in a permanently closed state, one sulfur atom is replaced by a carbon atom to generate an alternative bond that is locked in place under all biologically relevant conditions. In addition to inventing new chemical methods for exploring how disulfide bond status affects biological processes, this project directly impacts the training of undergraduate research students and also facilitates development of new engaging materials for the undergraduate science curriculum.Reactive oxygen/nitrogen species (ROS/RNS) are present in cells at various concentrations, depending on cell type, subcellular location, and physiological state. As an evolved response to the levels of ROS/RNS in oxidative metabolism, glutathione (GSH) is installed as a protected, but redox-reversible, covalent change to solvent-accessible cysteine thiols. By replacing the internal cysteine of GSH with an alkene-containing side chain, this modified GSH is irreversibly attached to solvent-accessible cysteines with optimized methods for thiol-ene coupling (TEC) in aqueous media. Initial targets include peroxiredoxins 1 & 2 (Prx1 & Prx2) and sulfiredoxin 1 (Srx1), where glutathionylation is critically involved in ROS/RNS equilibria and signaling. Mutagenesis of existing cysteine residues, followed by TEC reaction with the alkene-containing GSH, is assessed with LC-MS analysis, gel-filtration chromatography, spectrophotometry, analytical ultracentrifugation, and x-ray crystallography to explore how glutathionylation affects protein function and structure. With the developed TEC methodology, this project further contributes to the fundamental understanding of glutathionylation as a response to ROS/RNS.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

化学部的生命过程化学计划正在资助高点大学的Andrew J. Wommack博士研究蛋白质中硫-硫或二硫键的功能和结构后果。 二硫键在生物学中起着重要作用，从蛋白质的结构效应到细胞信号传导和代谢途径的控制。 这种硫-硫化学键可以在不同的生物条件下以打开或闭合状态存在，例如在不同的生理应激期间或在独特的细胞位置内。为了研究二硫键在永久闭合状态下的功能影响，将一个硫原子替换为一个碳原子，以生成一个替代键，该键在所有生物学相关条件下都被锁定在适当位置。除了发明新的化学方法来探索二硫键状态如何影响生物过程外，该项目还直接影响本科研究生的培训，并促进了本科科学课程新的引人入胜的材料的开发。活性氧/氮物质（ROS/RNS）以不同的浓度存在于细胞中，取决于细胞类型，亚细胞位置和生理状态。 作为对氧化代谢中ROS/RNS水平的进化反应，谷胱甘肽（GSH）被安装为受保护的，但氧化还原可逆的共价变化为溶剂可接近的半胱氨酸硫醇。 通过用含烯烃的侧链取代GSH的内部半胱氨酸，这种修饰的GSH用优化的方法在水介质中硫醇-烯偶联（TEC）不可逆地连接到溶剂可接近的半胱氨酸。最初的目标包括过氧化物酶1 - 2（Prx 1-Prx 2）和硫氧还蛋白1（Srx 1），其中谷胱甘肽化在ROS/RNS平衡和信号传导中起关键作用。现有的半胱氨酸残基的诱变，然后TEC反应与含烯烃的GSH，与LC-MS分析，凝胶过滤色谱，分光光度法，分析超离心和X-射线晶体学进行评估，探讨谷胱甘肽化如何影响蛋白质的功能和结构。 随着TEC方法的发展，该项目进一步促进了对谷胱甘肽作为ROS/RNS反应的基本理解。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。