Collaborative Research: Chemical Tools to Elucidate Glycolipid Biosynthesis and Transport in the Corynebacterineae

在化学系生命过程化学（CLP）项目的支持下，中密歇根大学的Benjamin Swarts教授和科罗拉多州立大学的玛丽杰克逊教授正在开发化学工具，以研究一类复杂的糖脂或糖连接的脂质分子，它们存在于数百种棒状杆菌亚目细菌的细胞被膜中。这些糖脂，被称为磷脂酰肌醇甘露糖苷（PIM），在细菌细胞防御，完整性和免疫活性中发挥重要作用。虽然PIM和相关分子是棒状杆菌科所特有的，并有助于这些生物体的独特的生理和致病特征，但这些糖脂如何在细菌细胞中构建和组织的许多方面仍然未知。这种知识差距仍然存在，因为使用传统技术研究PIM具有挑战性，并且缺乏分析它们的现代工具。该项目旨在开发新的化学生物学工具，以允许在细胞环境中对PIM和相关糖脂进行特定标记和跟踪。这个工具箱的应用程序可以提供一个更好的了解如何构建它们的细胞包膜。本科生和研究生以及从事这项研究的博士后研究员将获得有机合成，化学生物学和微生物学的跨学科培训。为了促进跨学科的培训和交流，两个机构的学生和博士后将定期举行虚拟会议，讨论该项目。该项目将被纳入一个推广计划，向社区大学生介绍化学细菌学研究，并鼓励他们继续接受STEM（科学，技术，工程和数学）教育。该项目中产生的工具预计将广泛适用，并支持其他科学家从事研究的Coryneacetylinositol mannosides（PIM）及其高度糖基化的衍生物，lipomannan（LM）和lipoarabinomannan（LAM），很难使用传统的生物化学和分子生物学技术进行研究，因为它们由高度复杂的，非遗传编码的脂质和糖组成。为了解决这个问题，该研究项目旨在开发基于合成碳水化合物的探针，这些探针利用天然和基因工程代谢途径将化学标签引入活细菌中的PIM，LM和LAM中。将在全细胞中测试荧光标记的糖脂分子，这可能提供一种在细胞环境中特异性追踪糖脂的方法。该项目中开发的探针有望提供以下信息：（i）活细菌细胞中PIM、LM和LAM构建的时空动力学;以及（ii）负责将PIM转运穿过细菌内膜的蛋白质的身份。该项目的研究结果有望提高对糖脂生物合成和动力学的理解，从而深入了解棒状杆菌亚目细菌的生理学。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估来支持。

With the support of the Chemistry of Life Processes (CLP) program in the Division of Chemistry, Professors Benjamin Swarts of Central Michigan University and Mary Jackson of Colorado State University are developing chemical tools to investigate a class of complex glycolipids, or sugar-linked lipid molecules, that are present in the cell envelope of hundreds of bacterial species from the suborder Corynebacterineae. These glycolipids, which are referred to as phosphatidylinositol mannosides (PIMs), play essential roles in bacterial cellular defense, integrity, and immunological activity. While PIMs and related molecules are unique to the Corynebacterineae and contribute to these organisms’ distinctive physiological and pathogenic characteristics, many aspects of how these glycolipids are constructed and organized in bacterial cells remain unknown. Such knowledge gaps remain because PIMs are challenging to study using traditional techniques and there is a lack of modern tools for analyzing them. This project seeks to develop new chemical biology tools to allow specific tagging and tracking of PIMs and related glycolipids in cellular contexts. Applications of this toolbox could provide a better understanding of how the Corynebacterineae construct their cell envelope. Undergraduate and graduate students and a postdoctoral fellow working on this research will acquire interdisciplinary training in organic synthesis, chemical biology, and microbiology. To promote interdisciplinary training and communication, students and postdocs at both institutions will meet regularly virtually to discuss the project. This project will be integrated into an outreach program to introduce community college students to chemical bacteriology research and to encourage their continued education in STEM (science, technology, engineering and mathematics). The tools generated in this project are expected to be broadly applicable and support other scientists pursuing research on the Corynebacterineae.Phosphatidylinositol mannosides (PIMs) and their highly glycosylated derivatives, lipomannan (LM) and lipoarabinomannan (LAM), are difficult to investigate using traditional techniques of biochemistry and molecular biology because they consist of highly complex, non-genetically encoded lipids and sugars. To address this problem, this research project seeks to develop synthetic carbohydrate-based probes that exploit native and genetically engineered metabolic pathways to introduce chemical tags into PIMs, LM, and LAM in live bacteria. Fluorescently labeled glycolipid molecules will be tested in whole cells, potentially providing a means to track the glycolipids specifically in a cellular context. The probes developed in this project are expected to provide information on (i) the spatiotemporal dynamics of PIM, LM, and LAM construction in live bacterial cells; and (ii) the identity of the protein(s) that is/are responsible for transport of PIMs across the bacterial inner membrane. The findings from this project are expected to provide an improved understanding of glycolipid biosynthesis and dynamics, which could provide insight into the physiology of bacteria in the Corynebacterineae suborder.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.