Exploiting Riboswitch Sensors to Reveal Antibiotics Uptake and Retention in Gram Negative Bacteria

利用核糖开关传感器揭示革兰氏阴性细菌中抗生素的摄取和保留

基本信息

批准号：
10343717
负责人：
RONALD R BREAKER
金额：
$ 104.76万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-02-19 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10343717
关键词：
Affect Anti-Bacterial Agents Antibiotic Therapy Antibiotics Bacteria Bacterial Physiology Binding Biochemical Biochemical Pathway Biochemistry Biological Process Cells Cellular Assay Chemicals Chemistry Data Development Dihydrofolate Reductase Inhibitor Enzymes Escherichia coli Exposure to Folic Acid Foundations Gene Expression Genes Gram-Negative Bacteria Growth Immunocompromised Host Infection Ions Knowledge Lead Libraries Ligands Measurable Mediating Messenger RNA Metabolic Pathway Metabolism Modeling Monitor Multi-Drug Resistance New Agents Organism Pathway interactions Penetration Permeability Pharmaceutical Chemistry Pharmacopoeias Physiological Processes Physiology Prevalence Pseudomonas aeruginosa RNA Regimen Reporter Reporter Genes Reporting Resistance Screening Result Signaling Molecule Structure-Activity Relationship System Systems Analysis Universities Untranslated RNA Validation absorption analog antimicrobial base cell envelope cheminformatics high throughput screening model development multidrug-resistant Pseudomonas aeruginosa next generation novel novel strategies opportunistic pathogen pathogen pharmacophore programs response screening sensor small molecule small molecule inhibitor small molecule libraries targeted agent therapeutic protein therapeutic target uptake

项目摘要

ABSTRACT Infections caused by MDR Pseudomonas aeruginosa and other Gram-negative pathogens challenge clinicians to find safe and effective antibiotic regimens that can eradicate these opportunistic pathogens from the frail, often immunocompromised hosts that they target. Two features of the P. aeruginosa cell envelope - its limited permeability to small molecules and the large number of both constitutive and inducible efflux systems it contains - render this pathogen intrinsically resistant to many available antimicrobials and contribute to acquired resistance toward the small set of existing anti-Pseudomonal antibiotics. This seriously limits the ability to identify “hits” with antibiotic activity using whole-cell assays - as compounds that penetrate and can inhibit key metabolic pathways are often effluxed out before they measurably inhibit bacterial growth or viability. Approaches that identify novel small molecule inhibitors of key bacterial enzymes often fail when these small molecules cannot achieve effective intrabacterial concentrations - and our understanding of the chemistries that would allow for penetration and retention is woefully incomplete. In this application we use a diverse array of riboswitches, sensitive and specific RNA-based small molecule sensors, as rapid and quantitative indicators that bacterial physiology has been perturbed. By multiplexing several riboswitches that report on accumulation of the alarmones ZTP and ppGpp, as well as the toxic product of increased SAM utilization, SAH, we can effectively screen for “signatures” of a bacterial response to sub- MIC levels of small molecules. Our approach places these riboswitch reporters in isogenic MDR and efflux- deficient P. aeruginosa strains, simultaneously yielding information about both physical and structural chemical features that allow penetration and efflux-avoidance and identifying “hit” molecules that can be developed as leads for new antibacterial agents. Our medicinal chemistry approach will build on both types of knowledge, allowing novel anti-Pseudomonal compounds to be identified and optimized.

抽象的由MDR假单胞菌和其他革兰氏阴性病原体挑战临床医生引起的感染为了找到可以从脆弱中放射这些机会性病原体的安全有效的抗生素方案，通常，他们针对的免疫功能低下的宿主。铜绿假单胞菌细胞信封的两个特征 - 有限对小分子的渗透性以及大量的本构和诱导的外排系统IT 包含 - 这种病原体对许多可用的抗菌剂具有本质上的抗性，并有助于对现有的一系列抗峰值抗生素产生了抵抗力。这严重限制了使用全细胞测定的抗生素活性识别“命中”的能力 - 作为穿透和可以的化合物抑制关键代谢途径通常会在测量抑制细菌生长或生存能力。鉴定新的小分子抑制剂的方法的方法在这些小分子无法获得有效的细菌浓度 - 我们对允许穿透和保留的化学条件严重不完整。在此应用中，我们使用了一系列核糖开关，敏感和特定的基于RNA的小分子传感器，作为细菌生理学的快速和定量指标。通过多路复用几种报告有关警报器ZTP和PPGPP的积累以及有毒产品的核糖开关 SAH的SAM利用率增加，我们可以有效筛选细菌反应的“特征” 小分子的麦克风水平。我们的方法将这些核糖开关记者置于等源性MDR和外排。缺乏铜绿假单胞菌菌株，仅产生有关物理和结构化学的信息允许渗透和避免外排的特征，并识别可以发展为可以发展为的“命中”分子新抗菌剂的铅。我们的医学化学方法将基于两种类型的知识，允许鉴定和优化新型的抗峰值化合物。

项目成果

期刊论文数量（16）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

The case of the missing allosteric ribozymes.

DOI：
10.1038/s41589-020-00713-2
发表时间：
2021-04
期刊：
Nature chemical biology
影响因子：
14.8
作者：
Panchapakesan SSS;Breaker RR
通讯作者：
Breaker RR

Comprehensive discovery of novel structured noncoding RNAs in 26 bacterial genomes.

DOI：
10.1080/15476286.2021.1917891
发表时间：
2021-12
期刊：
RNA biology
影响因子：
4.1
作者：
Brewer KI;Greenlee EB;Higgs G;Yu D;Mirihana Arachchilage G;Chen X;King N;White N;Breaker RR
通讯作者：
Breaker RR

Imaginary Ribozymes.

DOI：
10.1021/acschembio.0c00214
发表时间：
2020-08-21
期刊：
ACS chemical biology
影响因子：
4
作者：
Breaker RR
通讯作者：
Breaker RR

Architectures and complex functions of tandem riboswitches.

DOI：
10.1080/15476286.2022.2119017
发表时间：
2022-01
期刊：
RNA BIOLOGY
影响因子：
4.1
作者：
Sherlock, Madeline E.;Higgs, Gadareth;Yu, Diane;Widner, Danielle L.;White, Neil A.;Sudarsan, Narasimhan;Sadeeshkumar, Harini;Perkins, Kevin R.;Arachchilage, Gayan Mirihana;Malkowski, Sarah N.;King, Christopher G.;Harris, Kimberly A.;Gaffield, Glenn;Atilho, Ruben M.;Breaker, Ronald R.
通讯作者：
Breaker, Ronald R.

Employing a ZTP Riboswitch to Detect Bacterial Folate Biosynthesis Inhibitors in a Small Molecule High-Throughput Screen.