TrmD-targeting actinobacterial natural products as next generation antibiotics
TrmD靶向放线菌天然产物作为下一代抗生素
基本信息
- 批准号:10625857
- 负责人:
- 金额:$ 84.71万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2021
- 资助国家:美国
- 起止时间:2021-06-25 至 2026-05-31
- 项目状态:未结题
- 来源:
- 关键词:AccelerationAddressAnabolismAnti-Bacterial AgentsAntibioticsAnticodonBacteriaBindingBiochemistryBiological AssayBiologyBiotechnologyCell DeathCell FractionCellsCellular AssayClinicalCodon NucleotidesCollectionComplementComplexDoseDrug Binding SiteDrug TargetingEnzymesEscherichia coliExhibitsFluorescenceFundingGene ClusterGenesGram-Negative BacteriaGrowthHomo sapiensHumanInitiator CodonInvadedLibrariesMembraneMembrane ProteinsMessenger RNAMethodsMethylationMiningModelingModificationMolecularMulti-Drug ResistanceNatural ProductsNatural SelectionsNoisePenetrationPermeabilityPharmaceutical PreparationsPhenotypePopulationProductionProlineProtein BiosynthesisPublic HealthPumpReadinessReading FramesResearchResearch InstituteResistanceRibosomesRiskRisk AssessmentS-AdenosylhomocysteineS-AdenosylmethionineSideSignal TransductionSilverStructureTestingTimeTransfer RNATransferaseTranslationsbactericidecombatdrug discoveryefficacy studyefflux pumpgenome databasegenome sequencinghigh throughput screeningin vivomicrobialmortalitynext generationnovelpandemic diseaseposttranscriptionalprematureprogramsresistance mechanismresistance mutationresponsescaffoldscale upscreeningsmall molecule
项目摘要
Project Summary. Discovering new antibiotics for Gram-negative bacteria is uniquely challenging, due to their
double-membrane structure that acts as a permeability barrier to drugs and as an anchor for efflux pumps. Efforts
that target one membrane protein or one efflux pump at a time are ineffective, due to rapid rise of resistance
mutations. We will target the TrmD-catalyzed m1G37 methylation of tRNA to inhibit biosynthesis of multiple
classes of membrane proteins, with the potential to accelerate bactericidal action. TrmD is a bacteria-specific S-
adenosyl-methionine (AdoMet)-dependent methyl transferase that controls accuracy of the protein-synthesis
reading frame. Loss of TrmD increases +1 frameshifting and causes cell death. We have shown that genes for
multiple membrane proteins and efflux pumps in E. coli and in other Gram-negative bacteria contain TrmD-
dependent codons near the start of the reading frame. We hypothesize that targeting TrmD will reduce protein
synthesis of all of these genes, thus offering a novel solution to an unmet need. While AstraZeneca (AZ), GSK,
and academic labs have attempted to target TrmD by screening small molecular compound libraries, isolated
hits lack the cell-permeability needed to exhibit an antibacterial effect. Here, we propose to screen a large
collection of microbial extracts and fractions for cell-permeable and TrmD-targeting natural products (NPs) that
are potent and selective over the human counterpart Trm5. We will use a cell-based assay, consisting of a 1:1
mix of an E. coli (Ec) TrmDmCh strain (dependent on trmD for survival and expressing mCh (mCherry) as a
fluorescence marker) and an Ec Trm5YFP strain (dependent on trm5 for survival and expressing YFP), in a high-
throughput screening (HTS) campaign to isolate NPs that selectively inhibit the TrmDmCh strain. We perform this
assay in Ec tolC+ cells, which maintain the entire Gram-negative efflux machinery including the major efflux
pump encoded by tolC, to screen for NPs that are cell-permeable and resistant to efflux. A pilot screen with this
tolC+ cell-based assay has identified an attractive hit, demonstrating the HTS-readiness of the assay. In Aim 1,
we will use this tolC+ cell-based assay to screen 74,770 actinobacterial extracts and fractions available at The
Scripps Research Institute (TSRI). We will assess hits in secondary assays, remove false positives, evaluate
their activity at the whole-cell level, and test them for permeability and efflux in a panel of Gram-negative bacteria.
In Aim 2, we will de-replicate the top 20 hits to isolate the active NPs, determine their structures, and use a
combination of genome sequencing and mining to identify their biosynthetic gene clusters (BGCs) for developing
biotechnology platforms to scale up their production. In Aim 3, we will test active NPs for conferring TrmD-
deficient phenotypes in whole-cell assays, determine their potency, selectivity, mechanism of action, and assess
their risk of resistance. These NPs represent novel leads in a new paradigm of antibiotic discovery that addresses
the multi-drug resistance problem of Gram-negative bacteria.
项目总结。发现针对革兰氏阴性菌的新抗生素是一项独特的挑战,因为它们
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Ya-Ming Hou其他文献
Ya-Ming Hou的其他文献
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{{ truncateString('Ya-Ming Hou', 18)}}的其他基金
A cell model of YARS2-associated childhood-onset mitochondrial disease
YARS2 相关的儿童期发病线粒体疾病的细胞模型
- 批准号:
10575369 - 财政年份:2023
- 资助金额:
$ 84.71万 - 项目类别:
TrmD-targeting actinobacterial natural products as next generation antibiotics
TrmD靶向放线菌天然产物作为下一代抗生素
- 批准号:
10307014 - 财政年份:2021
- 资助金额:
$ 84.71万 - 项目类别:
TrmD-targeting actinobacterial natural products as next generation antibiotics
TrmD靶向放线菌天然产物作为下一代抗生素
- 批准号:
10438880 - 财政年份:2021
- 资助金额:
$ 84.71万 - 项目类别:
Exploring 3Dpol for RNA sequencing in real time
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- 批准号:
10166895 - 财政年份:2020
- 资助金额:
$ 84.71万 - 项目类别:
Exploring 3Dpol for RNA sequencing in real time
探索 3Dpol 实时 RNA 测序
- 批准号:
9974889 - 财政年份:2020
- 资助金额:
$ 84.71万 - 项目类别:
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