CAREER: Chemically-Enabled Strategies for the Discovery and Characterization of Novel Enzymatic Function

微生物不仅是地球上最丰富的生命形式，而且还具有最多样化的代谢功能。获得它们的集体化学武器库将大大提高我们生产从商品化学品到救生药品等基本小分子的能力。DNA测序技术的最新进展提供了丰富的微生物基因组，为发现新的酶提供了前所未有的机会。该项目的目标是识别这些基因组数据中编码的不寻常和潜在有用的酶化学。发现以前未表征的酶活性并将其引入我们的反应库将最终导致更有效和更环保的小分子合成途径。在这项工作中描述的特定的C-C键形成酶将是有机化学家和合成生物学家合成工具箱的重要补充。这些研究成果也将被整合到一个新的以实验室为基础的新生研讨会课程中，该课程将吸引学生从人类肠道微生物中发现新的酶。通过这个奖项，化学部门的生命过程化学项目资助了哈佛大学的Emily Balskus博士，以发现和表征在蓝藻天然产物的柱环烷家族的生物合成中介导不寻常的C-C键形成大环化二聚化步骤的酶。利用以化学知识为指导的基因组挖掘方法，PI发现了编码负责该反应的酶的生物合成基因簇。这些酶的功能和机制现在将通过体内和体外生化表征方法来阐明，这一努力将通过关键底物的化学合成来实现。通过与结构生物学家合作解决酶的晶体结构，也将获得重要的机制见解。该提案的中心教育目标是为一年级本科生创建一门新的研究型课程，重点是使用筛选方法从人类肠道微生物群中发现新的酶。本课程将把科学训练中最令人兴奋的方面带到大学课程的前沿，鼓励学生对进一步的本科研究和科学事业产生兴趣。总的来说，本提案中详细介绍的工作代表了化学和生物学界面上的综合研究和教育努力。

Microorganisms are not only the most abundant form of life on Earth, but they also possess the greatest diversity of metabolic functions. Gaining access to their collective chemical arsenal would significantly enhance our ability to produce essential small molecules ranging from commodity chemicals to life-saving pharmaceuticals. Recent advances in DNA sequencing technologies have delivered a wealth of microbial genomes that provide an unprecedented opportunity for the discovery of new enzymes. The goal of this project is to identify unusual and potentially useful enzymatic chemistry encoded in these genomic data. The discovery of previously uncharacterized enzymatic activity and its introduction into our reaction repertoire will ultimately lead to more efficient and environmentally friendly routes for small molecule synthesis. The specific C-C bond forming enzymes described in this work will be important additions to the synthetic toolkits of both organic chemists and synthetic biologists. These research efforts will also be integrated into a new laboratory-based freshman seminar course that will engage students in the discovery of new enzymes from human gut microbes. With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Emily Balskus from Harvard University to discover and characterize enzymes that mediate an unusual C-C bond-forming macrocyclizing dimerization step used in the biosynthesis of the cylindrocyclophane family of cyanobacterial natural products. Using a genome mining approach that is guided by chemical knowledge, the PI has uncovered the biosynthetic gene cluster encoding the enzymes responsible for this reaction. The functions and mechanisms of these enzymes will now be elucidated using in vivo and in vitro biochemical characterization methods, an effort that will be enabled by chemical synthesis of crucial substrates. Important mechanistic insights will also be obtained by solving the crystal structures of the enzymes in collaboration with structural biologists. The central educational objective of this proposal is to create a new research-based course for first-year undergraduates focused on using a screening approach to discover novel enzymes from the human gut microbiota. This class will bring the most exciting aspects of scientific training to the forefront of the college curriculum, encouraging interest in further undergraduate research and scientific careers. Overall, the work detailed in this proposal represents an integrated research and education effort at the interface of chemistry and biology.