RUI: Mechanistic Studies of the Firefly Luciferase Catalyzed Reactions

这个项目的目标是增加对生物如何将化学能转化为光的理解。生物发光，即生物有机体发出的光，是一种美丽的自然现象，它迷住了儿童，挑战了那些试图理解它的人，并为一种重要的研究工具提供了基础。生物发光的例子在自然界的细菌、蘑菇、水母、蚯蚓、文蛤、鱼和甲虫中都可以找到。该项目研究了萤火虫荧光素酶的结构与其在最终导致北美萤火虫产生黄绿光的两个连续反应的催化中的详细作用之间的关系。通过重组DNA技术制造的新的荧光素酶将通过光谱和动力学方法进行评估，以检查突变对每个反应的影响。这一方法也可能为荧光素酶结构中的基本构象变化的作用提供证据。这项研究的结果将增进对萤火虫生物发光的基本认识，以及生物将化学能转化为光的基本过程。该项目解决了生产新的荧光素酶蛋白的需要，以改善目前生物发光的研究应用，并推动新的生物发光蛋白的开发。本研究将在一所对本科生进行有效研究性训练的文科院校进行。现代化的设施和设备为学生和教职员工进行合作研究提供了良好的环境。这位首席研究员32年来一直积极指导本科生研究项目。约有85名本科生，其中大多数是妇女和少数族裔，曾与他共事，据估计，这些参与者中有70%进入了研究生或专业学校。该项目的目的是在多元文化环境中为不同的学生群体提供有意义的研究体验。该项目的所有学生和专业参与者将参与现代主流生物发光研究，并将为增加进入研究生课程和专业科学职业生涯的准备充分的毕业生数量做出积极贡献。这项研究的结果可能会改进萤火虫荧光素酶的用途，例如，在检测受细菌污染的食物中，或作为基因表达和调控的报告。

The goal of this project is to increase the understanding of how living organisms convert chemical energy into light. Bioluminescence, the emission of light by living organisms, is a beautiful natural phenomenon that has enchanted children, challenged those who have tried to understand it, and provided the basis for an important research tool. Examples of bioluminescence can be found throughout nature in bacteria, mushrooms, jellyfish, earthworms, clams, fish and beetles. This project addresses the relationship between the structure of the enzyme firefly luciferase and its detailed role in the catalysis of two sequential reactions that ultimately lead to the production of yellow-green light in the North American firefly, Photinus pyralis. New luciferases made by recombinant DNA techniques will be evaluated by spectroscopic and kinetics methods to examine the effects of mutations on each reaction. This approach may also provide evidence of the role of essential conformational changes in the luciferase structure. The results of this research will enhance the basic understanding of firefly bioluminescence and the fundamental process by which living organisms convert chemical energy into light. The project addresses the need to produce novel luciferase proteins to improve current research applications of bioluminescence and to advance the development of new ones.Broader Impacts. This research will be carried out at a liberal arts college with an established record of effective research training of undergraduate students. Modern facilities and equipment provide an excellent environment for students and faculty to engage in collaborative research. The principal investigator has actively directed undergraduate research projects for 32 years. Approximately 85 undergraduate students, the majority of whom are women and minorities, have worked with him, and an estimated 70% of these participants have entered graduate or professional school. The aim of this project is to offer a meaningful research experience to a diverse group of students in a multicultural environment. All student and professional participants in this project will be involved in modern mainstream bioluminescence research and will contribute positively to increasing the numbers of well-prepared graduates for entry into graduate programs and professional scientific careers. Results of this research may lead to improvements in the uses of firefly luciferase, for example, in the detection of food contaminated with bacteria or as a reporter of gene expression and regulation.