Characterization of a Family of Enzymes Responsible for the Modular Architecture of Nematode Pheromones

在化学系生命过程化学项目的支持下，来自佛罗里达大学的Rebecca Butcher将研究一个对线虫（蛔虫）中信号分子的产生至关重要的酶家族。 寄生线虫造成数十亿美元的农业损失，并感染世界上约三分之一的人口。 线虫通过分泌信号分子或信息素（也称为ascarosides）来相互交流，使蠕虫能够协调其发育以及各种行为，包括吸引、聚集和回避。 本研究计画将探讨羧酸酯酶（CEST）的酵素机制、底物及蛋白质结构。 这些酶负责介导影响其生物活性的子囊苷的特定化学修饰。 CEST酶的研究将有助于解剖线虫产生的大量信息素的生物学作用，并可能最终导致控制寄生线虫的新策略。 该项目还将通过开发基于课程的本科生研究体验（CURE）来增加STEM（科学，技术，工程和数学）专业的本科生保留率，为新生在本科生涯的早期提供真实的研究体验。 该研究项目将通过异源表达、纯化和表征其对候选底物的酶活性来表征关键CEST酶的生化活性。 与此同时，该项目将通过喂养蠕虫稳定的同位素标记的Ascarosides来表征体内CEST酶的底物范围。 将结晶代表性CEST酶，并使用X射线晶体学确定其结构，以研究其底物偏好的分子基础。 由于存在于蛔虫苷信息素的修改是至关重要的，他们的活动，这些修改是如何连接CEST酶的知识将有助于确定线虫信息素的生物学作用。 此外，CEST酶的酶活性的表征可以在未来使化学酶促合成的线虫信息素和衍生物库，这反过来又可以用来针对寄生nematodes.This奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估的支持。

With support from the Chemistry of Life Processes Program in the Division of Chemistry, Rebecca Butcher from the University of Florida will investigate a family of enzymes that are essential for the production of signaling molecules in nematodes (roundworms). Parasitic nematodes cause billions of dollars in agricultural damage and infect approximately one-third of the world’s human population. Nematodes communicate with each other by secreting signaling molecules or pheromones also known as ascarosides, that allow worms to coordinate their development, as well as various behaviors, including attraction, aggregation, and avoidance. This research project will study the enzymatic mechanisms, substrates, and protein structures of carboxylesterase (CEST) enzymes. These enzymes are responsible for mediating specific chemical modifications of ascarosides that influence their biological activities. Investigation of the CEST enzymes will be instrumental in dissecting the biological roles of the multitude of pheromones produced by nematodes and may ultimately lead to novel strategies for controlling parasitic nematodes. This project will also increase the retention of undergraduates in STEM (science, technology, engineering and mathematics) majors by developing a Course-based Undergraduate Research Experience (CURE) to provide freshmen with an authentic research experience early in their undergraduate careers. This research project will characterize the biochemical activities of key CEST enzymes by heterologously expressing them, purifying them, and characterizing their enzymatic activities against candidate substrates. In parallel, the project will characterize the substrate range of the CEST enzymes in vivo by feeding worms stable isotope-labeled ascarosides. Representative CEST enzymes will be crystallized, and their structures determined using X-ray crystallography in order to investigate the molecular basis for their substrate preferences. Because the modifications present in ascaroside pheromones are critical to their activities, knowledge of how these modifications are attached by CEST enzymes will help to determine the biological roles of nematode pheromones. Furthermore, characterization of the enzymatic activities of the CEST enzymes could in the future enable chemoenzymatic synthesis of nematode pheromones and libraries of derivatives, that could in turn be used to target parasitic nematodes.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.