Biosynthesis of Triplet States via Luciferase

通过荧光素酶生物合成三重态

• 批准号: 2109271
• 负责人: Michael Pirrung
• 金额: $ 47.4万
• 依托单位: University of California-Riverside
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2109271&HistoricalAwards=false
• 关键词: Biosynthesis; Triplet; States; via; Luciferase

With the support of the Chemistry of Life Processes (CLP) Program in the Division of Chemistry, Michael C Pirrung, Christopher J Bardeen both of the Department of Chemistry and Jiayu Liao of the Department of Bioengineering at the University of California-Riverside are studying the use of a bioluminescent system of the firefly to generate high energy states. Such states have unique chemical reactivity in biological systems and can convert ordinary oxygen into a species that kills cells. Because the system depends on a protein that can be genetically engineered, cell killing can be controlled as a dynamic response to conditions. This tool could, in principle, be exploited in synthetic biology and genetic logic circuits. Fundamental science will be advanced because no controlled means to generate such high energy states biologically is currently available. This project will benefit society via the provision of interdisciplinary training for students and the broadening of participation by groups underrepresented in science.This project will focus on using bioluminescence to efficiently generate triplet excited states, a process usually achieved for biological chromophores by photoexcitation. This research will fulfill a need for the controlled biological creation of singlet oxygen, a reactive oxygen species (ROS) involved in oxidative stress and the immune response. The investigators will achieve this goal by developing pairs of luciferase mutant proteins and synthetic luciferin analogs that can generate triplet states and, via sensitization, singlet oxygen, a species otherwise difficult to produce with biological / genetic control. Unnatural luciferin-luciferase pairs will be useful to modify biology by ablation of specific cells in vitro or in vivo based on gene expression of luciferase mutants. These goals will involve generation of non-native luciferase substrates, whose acceptance by wt firefly luciferase is typically much lower than luciferin. To maximize utility of these analogues, modification of the enzyme using protein engineering techniques will be undertaken.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.

在化学系生命过程化学（CLP）项目的支持下，加州大学河滨分校化学系的Michael C Pirrung，Christopher J Barrung和生物工程系的Jiayu Liao正在研究萤火虫生物发光系统的使用，以产生高能量状态。这种状态在生物系统中具有独特的化学反应性，可以将普通氧气转化为杀死细胞的物质。由于该系统依赖于一种可以通过基因工程改造的蛋白质，因此细胞杀伤可以作为对条件的动态响应来控制。原则上，这种工具可以用于合成生物学和遗传逻辑电路。基础科学将得到发展，因为目前还没有可控的方法来产生如此高的生物能量状态。本项目将通过为学生提供跨学科的培训，扩大科学领域代表性不足的群体的参与，造福社会。本项目将重点关注利用生物发光有效地产生三重激发态，这是生物发色团通常通过光激发实现的过程。这项研究将满足对单线态氧（一种参与氧化应激和免疫反应的活性氧（ROS））的受控生物产生的需求。研究人员将通过开发成对的荧光素酶突变蛋白和合成的荧光素类似物来实现这一目标，这些蛋白和类似物可以产生三重态，并通过敏化作用产生单线态氧，这是一种难以通过生物/遗传控制产生的物质。基于荧光素酶突变体的基因表达，非天然的荧光素酶-荧光素酶对可用于通过体外或体内消融特定细胞来修饰生物学。这些目标将涉及非天然荧光素酶底物的产生，其被wt萤火虫荧光素酶的接受度通常远低于荧光素。为了使这些类似物的效用最大化，将采用蛋白质工程技术对酶进行修饰。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。