Tool Compounds to Probe Multispecies Biofilms in the Human Oral Microbiome

用于探测人类口腔微生物组中多物种生物膜的工具化合物

• 批准号: 9533103
• 负责人: William M Wuest
• 金额: $ 37.01万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9533103
• 关键词: Affect; Bacteria; Binding; Biochemical; Biological; Biological Assay; Cells; Cellular Morphology; Characteristics; Chemicals; Childhood; Chronic; Clinical; Collaborations; Communities; Complex; Confocal Microscopy; Dental; Dental caries; Development; Disease; Drug Targeting; Endocarditis; Environment; Evaluation; Exhibits; Florida; Future; Genetic; Goals; Growth; Human; Investigation; Knock-out; Knowledge; Label; Laboratories; Lactams; Lead; Libraries; Light; Methods; Microbe; Microbial Biofilms; Microbiology; Molecular Genetics; Natural Products; Organic Synthesis; Organism; Phenotype; Process; Property; Proteomics; Research; Signal Transduction; Specificity; Streptococcus gordonii; Streptococcus mutans; Stress; Structure; Structure-Activity Relationship; Sum; Symbiosis; Synthesis Chemistry; System; Techniques; Testing; Therapeutic Agents; Work; analog; base; chemical stability; chemical synthesis; combat; commensal microbes; dental agent; design; drug development; experimental study; imaging probe; improved; in vivo; inhibitor/antagonist; interdisciplinary approach; interest; killings; nanomolar; oral bacteria; oral commensal; oral microbiome; oral pathogen; pathogen; pharmacophore; protein expression; response; scaffold; screening; tool; tool development; transcriptomics

Project Summary The proposed research will interrogate the natural product scaffold of carolacton as a starting point for agents that perturb S. mutans (SMU) biofilms. Carolacton is remarkable: instead of dispersing established biofilms or killing planktonic cells, carolacton targets cells transitioning into the biofilm state, and it is effective at very low concentrations. Here we present a multi-faceted approach, including organic synthesis, molecular genetics, proteomics, transcriptomics, and microbiological assays that begins with carolacton but has as an overarching goal the development of tool compounds that can be used to understand SMU biofilm processes within a multispecies environment. The first specific aim seeks to identify the specific target(s) of carolacton and a recently discovered analog that also has profound effects on SMU biofilms. This approach will employ genetic, transcriptomic, and MS-proteomic techniques to identify candidates that bind carolacton, from which we will confirm the targets using biochemical studies. This will shed light on the processes that the natural product affects and allow for a broader evaluation of the target in general biofilm processes. The second specific aim involves the chemical synthesis of rationally designed compounds related to carolacton. Proposed compounds will focus on four main aspects: 1) obtaining a detailed structure- activity relationship understanding of the carolacton structure; 2) determining the minimally complex scaffold that retains biological activity; 3) improving the physiochemical properties; and 4) identification of a lead compound for use in dental composites. Central to the efficient and concise strategies proposed is the knowledge gained in our previously described total synthesis. The third aim will investigate the biological properties of the tool compounds both in single species and multispecies biofilms. Preliminary results have identified that both carolacton and a newly synthesized analog cause a unique and specific phenotypic response to S. mutans biofilms. Therefore, all analogs will be tested against both planktonic and biofilm cells with S. mutans UA159 using confocal microscopy. Lead compounds will then be analyzed in assays with clinical isolates of both S. mutans and other bacteria involved in dental caries in collaboration with the Univ. of Rochester and the Univ. of Florida. Concurrently, compounds will be evaluated in mixed-species biofilms between S. mutans and commensal bacteria (i.e. S. sanguinis, S. gordonii) to identify pathogen-specific inhibitors. Previous work has demonstrated that oral pathogens with compromised signaling systems are reduced viability in the presence of commensal bacteria and we intend to build on this earlier work.

项目摘要 拟议的研究将审问卡洛克顿的天然产品支架作为起点 对于扰动S. mutans（SMU）生物膜的药物。卡洛克顿很棒：而不是分散 建立生物膜或杀死浮游细胞，卡洛克氏菌靶向过渡到生物膜状态的细胞， 它在非常低的浓度下有效。在这里，我们提出了一种多方面的方法，包括 有机合成，分子遗传学，蛋白质组学，转录组学和微生物学测定法 从卡洛克顿开始，但作为总体目标是工具化合物的开发 用于了解多物种环境中的SMU生物膜过程。 第一个特定目的旨在确定卡罗克顿的特定目标和最近发现的 对SMU生物膜也具有深远影响的类似物。这种方法将采用遗传， 转录组和MS-蛋白质组技术以识别结合Carolacton的候选者，从中 将使用生化研究确认靶标。这将阐明自然的过程 产品影响并允许对一般生物膜过程中的目标进行更广泛的评估。 第二个特定目的涉及与理性设计的化合物相关的化学合成 到卡罗克顿。建议的化合物将重点关注四个主要方面：1）获得详细的结构 - 卡洛克宫结构的活动关系理解； 2）确定最小复合物 保留生物活性的脚手架； 3）改善生理学特性； 4）识别 用于牙科复合材料的铅化合物。高效，简洁的策略的核心 提出的是我们先前描述的总合成中获得的知识。 第三个目标将研究单个物种中该工具化合物的生物学特性 和多物种生物膜。初步结果已经确定Carolacton和新的 合成的类似物对突变链球菌生物膜产生独特而特定的表型反应。所以， 所有类似物都将使用共焦点链球菌对浮游生物和生物膜细胞进行测试 显微镜。然后，将在两种螺旋链球菌的临床分离株中分析铅化合物 和其他与大学合作的细菌。罗切斯特和大学。的 佛罗里达。同时，将在S. utans和 共生细菌（即S. sanguinis，S。gordonii）鉴定病原体特异性抑制剂。以前的 工作表明，具有损害信号系统的口腔病原体可降低生存能力 在存在共生细菌的情况下，我们打算以早期的工作为基础。