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

合作研究：RUI：新型邻位氧螯合（VOC）双加氧酶L-DOPA双加氧酶的动力学研究和机制

• 批准号: 1708237
• 负责人: Keri Colabroy
• 金额: $ 18万
• 依托单位: Muhlenberg College
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1708237&HistoricalAwards=false
• 关键词: Collaborative; Research; RUI; Kinetic; Study

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.

有了这个奖项，化学部的生命过程化学项目资助了穆伦贝格学院的Keri Colabroy博士和罗兹学院的Larryn Peterson博士，以研究一种未研究的酶家族，这种酶家族可以切割称为儿茶酚的环状分子。儿茶酚环存在于植物的木质组织中，提供结构和强度。大自然将这些环改造成抗生素和其他生物活性材料。酶的结构和功能之间的复杂关系还不清楚。这种缺乏了解限制了利用酶来制造新的天然产品和将植物材料降解为生物燃料的能力。研究人员通过使用精心设计的小分子实时合作研究酶催化反应来获得新的知识，以揭示催化的内部工作原理。除了本科生在教师研究实验室的直接指导外，学生还可以在机构之间转移，从不同的角度体验项目。该项目的一部分被整合到每个家庭机构的本科课程中，以使更多的本科生有机会进行原创性研究。本研究中使用的现有和新兴技术和方法的教学视频发布到一个可免费访问的在线研究档案中，用于在合作机构内外培训下一代科学，技术，工程和数学劳动力。该研究项目通过探索结构和功能，扩展了对extradiol双加氧酶机制的理解，这些结构和功能是在邻近氧螯合物（VOC）超家族中的一个离散拓扑结构（IV型）组的代表性成员。新的小分子底物的合成燃料的反应动力学的研究，无论是在前稳态和平衡。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，并对底物和突变体数据进行全局动力学建模，以开发一种机制。这些实验拓宽了对extradiol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酶家族内拓扑结构的函数的双加氧酶机制的保守与置换的程度的机会。

项目成果

Derivatives of 3,4‐dihydroxyhydrocinnamic acid at the 6‐position as mechanistic probes of L‐DOPA dioxygenase

3,4-二羟基氢化肉桂酸 6-位衍生物作为左旋多巴双加氧酶的机械探针

• DOI: 10.1096/fasebj.2022.36.s1.r2554
• 发表时间: 2022
• 期刊: The FASEB Journal
• 作者: Steiner, Jessica;Xhafkollari, Gisella;Strzeminski, David J.;Leyes Porello, Sebastian;Colabroy, Keri L.;Peterson, Larryn W.
• 通讯作者: Peterson, Larryn W.

Extradiol cleavage of L‐DOPA as strategy for natural product biosynthesis

左旋多巴的额外二醇裂解作为天然产物生物合成的策略

• DOI: 10.1096/fasebj.2022.36.s1.r3491
• 发表时间: 2022
• 期刊: The FASEB Journal
• 作者: Jones, Paige A.;Johnson, Nyjah M.;Klugh, Kameron L.;Peterson, Larryn W.;Colabroy, Keri L.
• 通讯作者: Colabroy, Keri L.

Insight into L‐DOPA dioxygenase mechanism with 6‐substituted L‐DOPA derivatives

用6位取代的L-DOPA衍生物深入了解L-DOPA双加氧酶机制

• DOI: 10.1096/fasebj.2022.36.s1.r3477
• 发表时间: 2022
• 期刊: The FASEB Journal
• 作者: Nyamkondiwa, Kudzai;Squires, Trevor;Jones, Paige;Colabroy, Keri L.;Peterson, Larryn W.
• 通讯作者: Peterson, Larryn W.

A New Way of Belonging: Active-Site Investigation of L-DOPA Dioxygenase, a VOC Family Enzyme from Lincomycin Biosynthesis

一种新的归属方式：L-DOPA 双加氧酶（林可霉素生物合成中的 VOC 家族酶）的活性位点研究

• DOI: 10.1021/acs.biochem.9b00456
• 发表时间: 2019
• 期刊: Biochemistry
• 影响因子: 2.9
• 作者: Colabroy, Keri L.;Horwitz, Alyssa D.;Basciano, Victoria R.;Fu, Yizhi;Travitz, Kelly M.;Robinson, Miranda K.;Shimanski, Brittany A.;Hoffmann, Thomas W.
• 通讯作者: Hoffmann, Thomas W.

L‐DOPA dioxygenases from diverse natural product pathways

来自多种天然产物途径的左旋多巴双加氧酶

• DOI: 10.1096/fasebj.2022.36.s1.r3515
• 发表时间: 2022
• 期刊: The FASEB Journal
• 作者: Klugh, Kameron L.;Jones, Paige;Yoza, Riri;Peterson, Larryn W.;Colabroy, Keri L.
• 通讯作者: Colabroy, Keri L.