Mechanism and activity of beta-lactam resistant enzymes in E. faecium and E. faecalis
屎肠球菌和粪肠球菌中β-内酰胺抗性酶的机制和活性
基本信息
- 批准号:10624757
- 负责人:
- 金额:$ 70.12万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-05-09 至 2025-04-30
- 项目状态:未结题
- 来源:
- 关键词:AddressAffinityAmino Acid SubstitutionAntibiotic ResistanceAntibioticsArizonaBindingBiochemistryBiological AssayBiomolecular Nuclear Magnetic ResonanceCellular biologyCharacteristicsChemicalsChemistryClinicalCommunicable DiseasesCommunity-Acquired InfectionsComplexCoupledCrystallographyDangerousnessDataDevelopmentDoctor of PhilosophyEnterococcusEnterococcus faecalisEnterococcus faeciumEnzymesEssential Amino AcidsFamilyFutureGoalsHospitalsIn VitroInfectionInferiorKineticsKnowledgeLabelLipidsMapsMediatingMicrobiologyMolecularMutationN-terminalNMR SpectroscopyNosocomial InfectionsPenicillin ResistancePenicillin-Binding ProteinsPeptidoglycanPeptidyltransferasePredispositionProcessProtein DynamicsProteinsPublishingReagentRegulationResearch PersonnelResistanceRhode IslandRiceRoleSchemeStaphylococcus aureusStructureTestingTherapeuticTimeToxic effectTranslatingUniversitiesVariantanalogantimicrobialbasebeta-Lactam Resistancebeta-Lactamschemical synthesiscrosslinkdesignexperimental studyin vitro activityin vivoinsightinterdisciplinary approachmembermethicillin resistant Staphylococcus aureusmimeticsmultidisciplinarynovelpeptide chemical synthesisstructural biologysuicide substratessynergismtranspeptidation
项目摘要
Enterococci (e.g. E. faecalis and E. faecium) cause severe and often fatal nosocomial and community-acquired
infections. Therapy of enterococcal infections is frequently compromised by their decreased susceptibility
(increased resistance) to many classes of antibiotics, including β-lactams. This resistance is overwhelmingly
attributable to the expression of low-affinity penicillin-binding proteins PBP4 (E. faecalis) and PBP5 (E. faecium),
both of which are members of a family of low-affinity PBPs that also includes PBP2a from methicillin-resistant S.
aureus. In the clinical setting, E. faecium strains show widespread high-level penicillin resistance due to amino
acid substitutions, while similar highly-resistant E. faecalis strains are rare. Building on our extensive structural
and functional preliminary data, we will leverage the unique synergy of scientific expertise of the investigators to
answer the following key fundamental questions: how do low affinity PBPs bind and catalyze transpeptidation,
how do sequence changes in these PBPs further reduce their affinity for β-lactam antibiotics while retaining their
ability to synthesize peptidoglycan, and what cellular factors beyond low affinity PBP substitutions augment
levels of resistance expressed by clinical strains? To answer these questions, we will pursue four specific aims
that integrate structural biology, chemical synthesis, biochemistry and microbiology. Aim 1 will use structural
biology, especially biomolecular NMR spectroscopy, to determine why PBP5 is an inferior target of β-lactam
antibiotics. Our extensive preliminary data shows that this tour-de-force effort (at ~75 kDa, PBP5 is the second largest
single-chain protein studied using NMR spectroscopy) is not only feasible but, combined with our extensive
crystallographic data, will reveal why β-lactams only poorly inhibit PBP5 and, by extension, the entire family of low
affinity PBPs. Aims 2 and 3 will use newly developed chemical synthesis schemes coupled with structure
and dynamics (NMR spectroscopy) to determine how, at a molecular level, these PBPs catalyze
transpeptidation. We have achieved high-yield syntheses of PBP5-specific pentapeptide precursors and
variants of lipid II, enabling us to use NMR spectroscopy and transpeptidase assays to determine how substrates
bind and ultimately become cross-linked by PBP5. The impact of resistance-causing mutations in PBP5 on
transpeptidase activity will also be determined. Aim 4 will identify the orthogonal factors that contribute to
resistance in E. faecalis. Our preliminary data suggest that E. faecalis PBP2 likely contributes to β-lactam
resistance in the highly resistant LS4828 E. faecalis strain. We will quantify the contribution of PBP2 to LS4828
β-lactam resistance. In parallel, we will use BioID (proximity labeling) to identify PBP4 and PBP2 interacting
proteins (our recently published crystallographic data revealed that the PBP4 N-terminal domains are dynamic
and are likely involved in protein interactions). Together, these studies will reveal the structural and functional
details of enterococcal low-affinity PBP function, providing critical data upon which to base future strategies for
inhibiting these important enzymes.
肠球菌(如粪肠球菌和粪肠球菌)引起严重的,往往是致命的医院感染和社区感染
项目成果
期刊论文数量(3)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
Penicillin-Binding Proteins and Alternative Dual-Beta-Lactam Combinations for Serious Enterococcus faecalis Infections with Elevated Penicillin MICs.
青霉素结合蛋白和替代双 β-内酰胺组合治疗青霉素 MIC 升高的严重粪肠球菌感染。
- DOI:10.1128/aac.00871-22
- 发表时间:2023
- 期刊:
- 影响因子:4.9
- 作者:Cusumano,JaclynA;Daffinee,KathrynE;Ugalde-Silva,Paul;Peti,Wolfgang;Arthur,Michel;Desbonnet,Charlene;Rice,LouisB;LaPlante,KerryL;García-Solache,Mónica
- 通讯作者:García-Solache,Mónica
Enterococcal Physiology and Antimicrobial Resistance: The Streetlight Just Got a Little Brighter.
- DOI:10.1128/mbio.03511-20
- 发表时间:2021-02-23
- 期刊:
- 影响因子:6.4
- 作者:Rice LB
- 通讯作者:Rice LB
Molecular basis of β-lactam antibiotic resistance of ESKAPE bacterium E. faecium Penicillin Binding Protein PBP5.
- DOI:10.1038/s41467-023-39966-5
- 发表时间:2023-07-17
- 期刊:
- 影响因子:16.6
- 作者:Hunashal, Yamanappa;Kumar, Ganesan Senthil;Choy, Meng S.;D'Andrea, Everton D.;Da Silva Santiago, Andre;Schoenle, Marta V.;Desbonnet, Charlene;Arthur, Michel;Rice, Louis B.;Page, Rebecca;Peti, Wolfgang
- 通讯作者:Peti, Wolfgang
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{{ truncateString('Wolfgang Peti', 18)}}的其他基金
Serine/Threonine Phosphatases in Neurological Diseases
神经系统疾病中的丝氨酸/苏氨酸磷酸酶
- 批准号:
10583671 - 财政年份:2023
- 资助金额:
$ 70.12万 - 项目类别:
Shared Tundra screening cryo-EM for New England
新英格兰共享 Tundra 冷冻电镜筛查
- 批准号:
10413473 - 财政年份:2022
- 资助金额:
$ 70.12万 - 项目类别:
Protein Phosphatase 1 Holoenzyme Formation and Subunit Exchange
蛋白磷酸酶 1 全酶形成和亚基交换
- 批准号:
9985412 - 财政年份:2019
- 资助金额:
$ 70.12万 - 项目类别:
Mechanism and activity of beta-lactam resistant enzymes in E. faecium and E. faecalis
屎肠球菌和粪肠球菌中β-内酰胺抗性酶的机制和活性
- 批准号:
10391315 - 财政年份:2019
- 资助金额:
$ 70.12万 - 项目类别:
Mechanism and activity of beta-lactam resistant enzymes in E. faecium and E. faecalis
屎肠球菌和粪肠球菌β-内酰胺抗性酶的机制和活性
- 批准号:
9927573 - 财政年份:2019
- 资助金额:
$ 70.12万 - 项目类别:
Dynamics & energetics of p38a kinase regulation by ligands
动力学
- 批准号:
8608555 - 财政年份:2013
- 资助金额:
$ 70.12万 - 项目类别:
Dynamics & energetics of p38a kinase regulation by ligands
动力学
- 批准号:
8436569 - 财政年份:2013
- 资助金额:
$ 70.12万 - 项目类别:
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