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.

抽象的耐多药铜绿假单胞菌和其他革兰氏阴性病原体引起的感染给人群带来挑战找到安全有效的抗生素治疗方案，可以根除体弱者身上的这些机会性病原体，铜绿假单胞菌细胞包膜的两个特征——其局限性。对小分子的渗透性以及大量的组成型和诱导型外排系统含有 - 使这种病原体对许多可用的抗菌药物具有内在的抗药性，并有助于对现有的一小部分抗假单胞菌抗生素产生了耐药性，这严重限制了它的应用。能够使用全细胞检测来识别具有抗生素活性的“目标”——因为化合物可以渗透并可以抑制关键代谢途径的物质通常在明显抑制细菌生长或识别关键细菌酶的新型小分子抑制剂的方法通常会失败。这些小分子无法达到有效的细菌内浓度——我们对这些小分子的理解令人遗憾的是，允许渗透和保留的化学物质是不完整的。在此应用中，我们使用多种核糖开关、敏感且特异的基于 RNA 的小分子传感器，作为细菌生理学受到干扰的快速定量指标。几个报告报警素 ZTP 和 ppGpp 以及有毒产物积累的核糖开关随着 SAM 利用率的增加，SAH，我们可以有效地筛选细菌对亚细菌反应的“特征” 我们的方法将这些核糖开关生产者置于同基因 MDR 和外排中。缺陷型铜绿假单胞菌菌株，同时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，并识别可开发为的“命中”分子我们的药物化学方法将建立在这两类知识的基础上，允许识别和优化新型抗假单胞菌化合物。

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.