Genetically-encoded fluorescent RNA sensors for measuring transport of antibiotics into the cytoplasm of Gram-negative pathogens and development of efflux pump inhibitors

用于测量抗生素转运至革兰氏阴性病原体细胞质的基因编码荧光RNA传感器以及外排泵抑制剂的开发

• 批准号: 10326785
• 负责人: STEPHEN LORY
• 金额: $ 60.52万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-02-13 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10326785
• 关键词: Affinity; Animals; Antibiotics; Assessment tool; Bacteria; Binding; Biological; Biological Assay; Broccoli - dietary; Catalytic RNA; Cells; Chemicals; Chimeric Proteins; Clinical; Collection; Combined Modality Therapy; Communities; Crystallization; Cytoplasm; DNA; Detection; Development; Diagnostic; Disease; Drug Design; Drug Industry; ESKAPE pathogens; Effectiveness; Engineering; Family; Flow Cytometry; Goals; Human; Individual; Industrialization; Infection; Laboratories; Libraries; Life; Measures; Mediating; Medical; Membrane; Membrane Fusion; Membrane Proteins; Microscopy; Modeling; Multi-Drug Resistance; Nodule; Organism; Pathogenicity; Penetration; Performance; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phase; Phenotype; Property; Proteins; Pseudomonas aeruginosa; Public Health; Pump; RNA; RNA Binding; Reporter; Research; Research Personnel; Resistance; Series; Structure; Testing; Therapeutic; Therapeutic Agents; aptamer; bactericide; base; cell envelope; combat; combinatorial; detector; diagnostic tool; drug discovery; efflux pump; improved; in vivo; inhibitor; member; microorganism; novel; novel therapeutics; pathogen; periplasm; prevent; response; screening; screening program; sensor; small molecule; tool; uptake

Project Summary Complications from infections caused by difficult to treat Gram-negative pathogens resistant to multiple antibiotics (the MDR phenotype) has become a major public health threat. The basis of resistance in the most important members of this group, including Pseudomonas aeruginosa, is the poor penetration of the antibiotics through a relatively impermeable bacterial cell envelope and their expulsion (efflux) by various pumps expressed by the microorganisms. Consequently, the intracellular concentrations of antibiotic are kept below bactericidal levels. This proposal aims at creating new classes of diagnostic tools and therapeutic agents to combat efflux-based resistance by a project organized into two phases. First, we will develop genetically encoded fluorescent sensors activated by binding of antibiotics following their entry into the bacterial cytoplasm. This tool will enable us to measure antibiotic flux under different environmental conditions and on a single cell basis. During the second phase of the project, working with our industrial partner, we will implement a screening program, using a library of several hundred billion small molecules, to identify inhibitors targeting different components of efflux pumps. These inhibitors should activate the fluorescent detector by causing enhanced intracellular accumulation of the antibiotics resulting from inhibition of efflux. Functional studies are proposed to assess the ability of the active compounds to promote killing of P. aeruginosa by antibiotics that are normally expelled by the efflux pumps. Crystal structures of inhibitors bound to target proteins will be determined and these will guide medicinal chemistry efforts with the goal of creating additional derivatives with increased potencies and expanded spectrum, capable of inhibiting efflux pumps of other important Gram-negative pathogens.

项目总结

项目成果

Rational Design of Allosteric Fluorogenic RNA Sensors for Cellular Imaging.

用于细胞成像的变构荧光 RNA 传感器的合理设计。

• DOI: 10.1007/978-1-0716-1499-0_11
• 发表时间: 2021
• 期刊: Methods in molecular biology (Clifton, N.J.)
• 作者: Yu,Qikun;Zheng,Ru;Narayanan,Manojkumar;You,Mingxu
• 通讯作者: You,Mingxu

The core and accessory Hfq interactomes across Pseudomonas aeruginosa lineages.

• DOI: 10.1038/s41467-022-28849-w
• 发表时间: 2022-03-10
• 期刊: Nature communications
• 影响因子: 16.6
• 作者: Trouillon J;Han K;Attrée I;Lory S
• 通讯作者: Lory S

Paper-based fluorogenic RNA aptamer sensors for label-free detection of small molecules.

基于纸张的荧光RNA适体传感器，用于无标记的小分子。

• DOI: 10.1039/d0ay00588f
• 发表时间: 2020-06-04
• 期刊: Analytical methods : advancing methods and applications
• 作者: Shafiei F;McAuliffe K;Bagheri Y;Sun Z;Yu Q;Wu R;You M
• 通讯作者: You M

A Genetically Encoded RNA Photosensitizer for Targeted Cell Regulation.

用于靶向细胞调节的遗传编码的RNA光敏剂。

• DOI: 10.1002/anie.202010106
• 发表时间: 2020-12-01
• 期刊: Angewandte Chemie (International ed. in English)
• 作者: Ren K;Keshri P;Wu R;Sun Z;Yu Q;Tian Q;Zhao B;Bagheri Y;Xie Y;You M
• 通讯作者: You M

Toward a Comprehensive Analysis of Posttranscriptional Regulatory Networks: a New Tool for the Identification of Small RNA Regulators of Specific mRNAs.

• DOI: 10.1128/mbio.03608-20
• 发表时间: 2021-02-23
• 期刊: mBio
• 影响因子: 6.4
• 作者: Han K;Lory S
• 通讯作者: Lory S