Mulitvalent Ligands as Effectors

作为效应器的多价配体

• 批准号: 7099215
• 负责人: Laura L Kiessling
• 金额: $ 29.63万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-06-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7099215
• 关键词: Bacillus subtilis; Escherichia coli; G protein; Proteus; binding proteins; biofilm; biological signal transduction; cell differentiation; chemoattractants; chemokine; chemoreceptors; chemotaxis; gene expression profiling; ions; leukocytes; ligands; method development; peptide library; protein protein interaction; protein structure function; receptor binding

DESCRIPTION (provided by applicant): This proposal is a competitive renewal of program with a long-term objective to elucidate the molecular details of chemotactic signaling in bacteria using chemical biology strategies. A second objective of the proposed research is to ascertain whether our findings in bacteria are relevant for leukocyte chemotaxis. The signaling pathway that results in bacterial chemotaxis serves as a model for understanding two- component signaling in particular and transmembrane signaling in general. Bacteria respond sensitively to changes in chemoeffector concentration over a large concentration range. We hypothesize that bacteria amplify subtle differences in signal concentration through a signaling lattice, and there is mounting evidence for such a model. This model raises several questions: How are proteins organized within the lattice? How does attractant binding alter interactions within the lattice? What is the effect of repellents on the lattice structure? How does covalent modification (phosphorylation and methylation) affect interactions within the lattice? Are there parallels in the signal amplification mechanisms of bacteria and neutrophils? We propose to address these questions using a variety of approaches. These include the synthesis and testing of monovalent and multivalent chemoeffectors, protein footprinting experiments, and x-ray crystallography. Aim 1 is focused on understanding how periplasmic binding proteins (PBPs) function. We have identified a ligand for the periplasmic glucose/galactose binding protein (GGBP), and this ligand inhibits GGBP- mediated chemotaxis. We propose to carry out structural studies to understand how this PBP antagonist functions. We envision that such studies will be useful in devising inhibitors of other bacterial processes mediated by PBPs including biofilm formation and virulence. In aim 2, we plan to continue studies initiated in the past grant period explore the role of inter-chemoreceptor interactions in chemotactic signal amplification in bacteria. In aim 3, we propose to use protein footprinting experiments to investigate protein - protein interactions important in chemotaxis signal transduction. We anticipate that the studies proposed in aims 2 and 3 will provide an increased understanding of how bacteria transduce and amplify signals. In aim 4, we propose to evaluate multivalent ligands as leukocyte chemoattractants. The goal of this aim is to determine whether there are similarities in the mechanisms by which bacteria and leukocytes amplify signals. We anticipate that our studies will facilitate the development of agents that interfere with key two- component signaling pathways; therefore, they may result in the development of new anti-microbial agents. Moreover, neutrophil chemotaxis occurs in the inflammatory/immune response. Thus, conclusions from our studies will be broadly applicable to understanding other signaling pathways involved in human disease.

描述（由申请人提供）：该提案是对程序的竞争续订，其长期目标是使用化学生物学策略阐明细菌中趋化信号的分子细节。拟议研究的第二个目标是确定我们在细菌中的发现是否与白细胞趋化性有关。 导致细菌趋化性的信号传导途径是一种模型，尤其是理解两个组件信号的模型，尤其是跨膜信号传导。细菌对在较大浓度范围内化学作用者浓度的变化敏感。我们假设细菌通过信号晶格扩大了信号浓度的细微差异，并且有越来越多的证据证明了这种模型。该模型提出了几个问题：如何在晶格中组织蛋白质？吸引人的结合如何改变晶格内的相互作用？驱虫剂对晶格结构有什么影响？共价修饰（磷酸化和甲基化）如何影响晶格内的相互作用？细菌和中性粒细胞的信号扩增机制中是否存在相似之处？我们建议使用各种方法解决这些问题。这些包括单价和多价Chemoefector的合成和测试，蛋白质足迹实验和X射线晶体学。 AIM 1的重点是了解周质结合蛋白（PBP）的功能。我们已经鉴定出用于周质葡萄糖/半乳糖结合蛋白（GGBP）的配体，该配体抑制了GGBP-介导的趋化性。我们建议进行结构研究，以了解该PBP拮抗剂的功能。我们设想，此类研究将有助于设计由PBP介导的其他细菌过程的抑制剂，包括生物膜形成和毒力。在AIM 2中，我们计划继续在过去的赠款期间开始研究，探讨了细菌中受体感受人相互作用在趋化信号扩增中的作用。在AIM 3中，我们建议使用蛋白质足迹实验来研究在趋化性信号转导中重要的蛋白质 - 蛋白质相互作用。我们预计AIM 2和3中提出的研究将对细菌如何转导和扩增信号的方式有更多的了解。在AIM 4中，我们建议评估多价配体作为白细胞趋化剂。该目标的目的是确定细菌和白细胞扩增信号的机制中是否存在相似之处。 我们预计我们的研究将有助于干扰关键两分组件信号通路的代理的发展。因此，它们可能导致新的抗微生物剂的发展。此外，中性粒细胞趋化性发生在炎症/免疫反应中。因此，我们的研究得出的结论将广泛适用于理解与人类疾病有关的其他信号通路。