Collaborative Research: RUI: Kinetic Study and Mechanism of L-DOPA dioxygenase, a new type of vicinal-oxygen-chelate (VOC) dioxygenase

有了这个奖项，化学学部的生命过程化学项目资助了来自穆伦伯格学院的Keri Colabroy博士和罗德学院的Larryn Peterson博士来研究一种尚未被研究的酶家族，这种酶可以切割被称为儿茶酚的环状分子。在植物的木质组织中发现了天主教环，以提供结构和力量。大自然将这些环重新塑造成抗生素和其他生物活性物质。酶的结构与功能之间的复杂关系尚不清楚。这种认识的缺乏限制了利用酶来制造新的天然产物和将植物材料降解为生物燃料的能力。研究人员通过对酶催化反应的实时合作研究产生了新的知识，使用精心设计的小分子来揭示催化的内部工作原理。除了在学院研究实验室直接指导本科生之外，学生们还可以在不同的机构之间转学，从不同的角度体验项目。项目的一部分被整合到每个本土院校的本科课程中，让更多的本科生有机会进行原创研究。本研究中使用的现有和新兴技术和方法的教学视频将发布到一个免费访问的在线研究档案中，用于培训合作机构内外的下一代科学、技术、工程和数学工作人员。本研究项目通过探索邻近氧螯合物（VOC）超家族中一个离散拓扑（IV型）组的结构和功能，扩展了对外二醇双加氧酶机制的理解。新型小分子底物的合成促进了反应动力学的研究，无论是在预稳定状态还是平衡状态。这项工作还使用x射线晶体学和诱变来定义活性位点残基的作用，并使用底物和突变体数据的全局动力学建模来开发机制。这些实验拓宽了对外二醇双加氧酶机制的结构和动力学理解，同时也提供了了解作为VOC酶家族拓扑功能的双加氧酶机制的保护程度和排列程度的机会。

With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Keri Colabroy from Muhlenberg College and Dr. Larryn Peterson of Rhodes College to investigate an unstudied family of enzymes that cleave ring-shaped molecules called catechols. Catecholic rings are found in the woody tissue of plants to provide structure and strength. Nature refashions these rings into antibiotics and other bioactive materials. The complex relationship between enzyme structure and function is not well understood. This lack of understanding limits the ability to exploit the use of enzymes to make new natural products and degrade plant material into biofuels. The researchers generate new knowledge through the collaborative study of the enzyme-catalyzed reactions in real-time using small molecules carefully designed to expose the inner workings of catalysis. In addition to the direct mentorship of undergraduate students in faculty research labs, students transfer between institutions to experience the project from different perspectives. Part of the project is integrated into undergraduate coursework at each of the home institutions to give additional undergraduates the opportunity to conduct original research. Instructional videos of the established and emerging technologies and methods used in this research are published to a freely-accessible, online research archive for training of the next-generation science, technology, engineering and mathematics workforce both at the partner institutions and beyond. This research project expands understanding of the extradiol dioxygenase mechanism by exploring structure and function for evolutionarily distinct, but representative members of a discreet topology (type IV) group within the vicinal-oxygen-chelate (VOC) superfamily. The synthesis of novel small-molecule substrates fuel the study of reaction kinetics, both in the pre-steady state and at equilibrium. This work also uses X-ray crystallography and mutagenesis to define the roles of active site residues and global kinetic modeling of substrate and mutant data to develop a mechanism. These experiments broaden the structural and kinetic understanding of extradiol dioxygenase mechanism, while also providing the opportunity to understand the extent of conservation versus permutation of dioxygenase mechanism as a function of topology within the VOC enzyme family.