MIRA: Probing Glycan Polymer Patterns on Bacterial Cell Surfaces

项目总结/摘要 我们的星球上居住着数万亿的细菌，它们生活在人类的体内和体外。“皮肤”，或 细菌的表面被称为细胞被膜，其功能是将我们与它们分开。虽然有些细菌 虽然细菌是共生体，但病原菌感染仍然是全世界的主要死因。而革兰氏阴性 细菌含有保护性的外膜层，大多数革兰氏阳性菌都没有，几乎所有的细菌都含有 由从细胞表面延伸的独特聚糖模式组成的聚合物。细菌表面糖 聚合物，或外多糖，作为分子条形码，区分不同菌株的细菌内， 一个单一的物种许多细菌胞外多糖含有稀有糖，这些糖是单糖， 包括人类在内的其他生物体中不存在。虽然胞外多糖是宿主感染所必需的，但我们 仍然缺乏对罕见的含糖聚糖聚合物如何组装、识别和使 在宿主体内生存。 我的实验室寻求产生化学和生物化学工具来研究细菌蛋白质和聚糖 使其能够在不同的环境中生存。我们的主要重点领域是：（1）发展小型 细菌伴侣功能的分子调节剂;（2）操纵细胞表面糖模式， 标记并使细菌失效。这项建议的重点是后一个程序，其中我们确定罕见的糖 称为O抗原的革兰氏阴性细胞表面聚合物所特有的亚基，代表关键表位 介导与宿主的相互作用和对抗生素的敏感性。在未来五年，我们将致力于 以下问题：（1）我们能否改进化学酶促途径，以稀有糖前体底物？(2)如何 糖基转移酶识别稀有糖底物来构建O抗原吗？(3)是O抗原 通过蛋白质-蛋白质相互作用调节的糖基转移酶？(4)宿主蛋白质的结构基序是什么 参与了细菌对稀有糖的识别(5)我们能否鉴定出与细菌感染有关的新的宿主蛋白质 认可？为了回答这些问题，我们将采用多学科的方法，包括 有机化学、化学生物学、生物化学、微生物学和测序分析。这项工作将 显著扩展了我们对细菌多糖合成细胞机制的理解， 并将教会我们人类如何识别外来糖。 与公共卫生的相关性：除了提供对细菌因子产生的基本见解外， 这一提议的结果将为新的策略提供信息， 革兰氏阴性菌感染的干扰基本宿主-病原体相互作用，以及生物分子 用于识别细菌寡糖结构的新诊断试剂。

Project Summary/Abstract Our planet is inhabited by trillions of bacteria that live inside and outside of humans. The “skin”, or surface, of bacteria is called the cell envelope, and functions to separate us from them. Although some bacteria are symbionts, infection by pathogenic bacteria is still a major cause of death worldwide. While Gram-negative bacteria contain a protective outer membrane layer absent in most Gram-positives, almost all bacteria contain polymers composed of unique patterns of glycans that extend from the cell surface. Bacterial surface sugar polymers, or exo-polysaccharides, act as molecular barcodes that distinguish different strains of bacteria within a single species. Many bacterial exo-polysaccharides contain rare sugars, which are monosaccharides that are absent in other organisms, including humans. While exo-polysaccharides are necessary for host infection, we still lack an understanding of how rare sugar-containing glycan polymers are assembled, recognized, and enable survival in the host. My laboratory seeks to generate chemical and biochemical tools to study bacterial protein and glycan pathways that enable survival in different environments. Our main areas of focus are: (1) development of small molecule regulators of bacterial chaperone function; (2) manipulation of cell surface sugar patterns to selectively label and disable bacteria. This proposal focuses on the latter program, in which we identify rare saccharide subunits that are unique to Gram-negative cell surface polymers called O-antigens, and represent key epitopes that mediate interactions with hosts and susceptibility to antibiotics. Over the next five years, we will address the following questions: (1) Can we improve chemoenzymatic routes to rare sugar precursor substrates? (2) How do glycosyltransferases recognize rare sugar substrates to build O-antigens? (3) Are O-antigen glycosyltransferases regulated via protein-protein interactions? (4) What host protein structural motifs are involved in bacterial rare sugar recognition? (5) Can we identify new host proteins involved in bacterial recognition? To answer these questions, we will use a multidisciplinary approach, involving a combination of organic chemistry, chemical biology, biochemistry, microbiology and sequencing-based analyses. This work will significantly expand our understanding of cellular mechanisms underlying bacterial polysaccharide synthesis, and will teach us how humans recognize foreign sugars. Relevance to public health: In addition to providing fundamental insight into the production of bacterial factors that are important for infection, the results of this proposal will inform novel strategies to disable hard-to-treat Gram-negative infections by interference of essential host-pathogen interactions, as well as biomolecular reagents to recognize bacterial oligosaccharide structures for new diagnostics.