Ultra-sensitive NMR via Photochemically induced dynamic nuclear polarization
通过光化学诱导动态核极化的超灵敏核磁共振
基本信息
- 批准号:7991252
- 负责人:
- 金额:$ 23.5万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2010
- 资助国家:美国
- 起止时间:2010-06-03 至 2012-05-31
- 项目状态:已结题
- 来源:
- 关键词:Amino AcidsAntibioticsApplications GrantsAreaArgonAromatic Amino AcidsAromatic CompoundsBindingBiologicalBiological ProcessBiologyCationsCell NucleusCellsChemicalsChemistryComplexComputer softwareDetectionDevelopmentDiffusionDisadvantagedElectronsEngineeringEnvironmentEscherichia coliExhibitsFlavinsGasesGoalsHandIndolesInvestigationIonsLasersLightLiquid substanceMedicineMethodsModelingMolecularMolecular ChaperonesNMR SpectroscopyNational Institute of Allergy and Infectious DiseaseNuclearNuclear Magnetic ResonancePhysiologic pulsePrincipal InvestigatorProteinsResearchResearch ProposalsResolutionRoleSamplingScienceShapesSideSimulateSolidSolutionsSolventsTechniquesTemperatureTestingTranslationsUncertaintyViscosityanalogbasecontextual factorsdensityelectron densityexperienceimprovedinterestmethod developmentnovelnovel strategiespolypeptideprotein foldingpublic health relevanceresearch studytool
项目摘要
DESCRIPTION (provided by applicant): NMR spectroscopy suffers from an extremely serious disadvantage: very low sensitivity. The goal of this project is to develop novel approaches to enhance the sensitivity of liquid state nuclear magnetic resonance (NMR) and optimize the detection limits of biomolecules containing aromatic amino acids. This research will take advantage of method developments in heteronuclear correlation photo-chemically-induced dynamic nuclear polarization, also known as photo-CIDNP, and it will employ NMR setups in conjunction with an Argon Ion laser able to photo-excite flavin-containing NMR samples at 488 nm. The following three specific research aims will be pursued. Specific Aim #1: Development of novel 13C-based photo-CIDNP pulse sequences involving 13C-1H heteronuclear correlation. Several novel 2D pulse sequences will be developed and tested on both free aromatic amino acids (including Trp, which exhibits a strong 1H photo-CIDNP) and aromatic amino acids belonging to the model protein apoHmpH. The effect of laser power (up to 25W), laser duration, temperature, and viscosity will also be systematically explored. We perceive this aim as extremely exciting, in light of our recent results on 15N1H heteronuclear correlation and preliminary ab initio calculations, which strongly support the feasibility of this approach. Specific Aim #2: Ab initio electron density calculations on model substituted indoles and other aromatic compounds resembling the side chain of naturally occurring amino acids and amino acid analogs. These calculations will be based on restricted Hartree-Fock and Density Functional approaches. The computations will be carried out with the Gaussian software package, available in the UW-Madison Department of Chemistry. The goal of this aim is to evaluate the hyperfine couplings of radical cations involved in photo-CIDNP, for the relevant nuclei of interest. The results of these studies will inform the method developments in Specific Aim #1. Specific Aim #3: Application of the methods developed in Specific Aim #1 to study the binding of polypeptides rich in aromatic amino acids to the bacterial trigger factor (TF) molecular chaperone at low concentrations. In addition to the TF from E. coli, an aromatic-rich polypeptide derived from the Hmp protein (comprising its residues 116-134) will be utilized for these studies. The proposed studies are aiming at pushing the sensitivity boundaries of liquid state NMR under mild, physiologically relevant conditions suitable for biological applications in dilute solutions.
PUBLIC HEALTH RELEVANCE: This project targets novel approaches to enhance the sensitivity of liquid state nuclear magnetic resonance (NMR) and optimize the detection limits of biomolecules containing aromatic amino acids by photo-chemically- induced dynamic nuclear polarization (photo-CIDNP). Given the applicability of this approach to both aromatic amino acids within proteins and aromatic-ring-containing antibiotics, this research has direct implications for the sensitive detection of antibiotic binding to their biomolecular targets, one of the aims of NIAID. The proposed applications to the bacterial chaperone trigger factor (TF) are relevant to the bacterial translation machinery, a frequent target for antibiotic development.
描述(由申请人提供):核磁共振波谱有一个非常严重的缺点:非常低的灵敏度。本项目的目标是开发新的方法来提高液态核磁共振(NMR)的灵敏度,并优化含有芳香氨基酸的生物分子的检测限。本研究将利用异核相关光化学诱导动态核极化(也称为光- cidnp)方法的发展,并将核磁共振装置与能够在488 nm光激发含黄素的核磁共振样品的氩离子激光器结合使用。以下三个具体的研究目标将被追求。具体目标#1:开发新的基于13C-1H异核相关的光- cidnp脉冲序列。将开发几种新的二维脉冲序列,并对游离芳香氨基酸(包括表现出强1H光cidnp的色氨酸)和属于模型蛋白apoHmpH的芳香氨基酸进行测试。还将系统地探讨激光功率(最高可达25W)、激光持续时间、温度和粘度的影响。鉴于我们最近在15N1H异核相关和初步从头计算方面的结果,我们认为这一目标非常令人兴奋,这有力地支持了这种方法的可行性。具体目标#2:从头计算模型取代吲哚和其他芳香族化合物的电子密度,这些化合物类似于天然存在的氨基酸和氨基酸类似物的侧链。这些计算将基于受限的Hartree-Fock和密度泛函方法。计算将进行高斯软件包,可在化学系威斯康星大学麦迪逊分校。本目的目的是评估参与光cidnp的自由基阳离子的超精细耦合,以获得相关的感兴趣的原子核。这些研究的结果将为具体目标#1中的方法发展提供信息。特异性目标#3:应用特异性目标#1中开发的方法,研究富含芳香氨基酸的多肽在低浓度下与细菌触发因子(TF)分子伴侣的结合。除了来自大肠杆菌的TF外,从Hmp蛋白(包括其残基116-134)衍生的富含芳香的多肽将用于这些研究。拟议的研究旨在推动液态核磁共振在温和的、生理相关的条件下的灵敏度边界,适用于稀溶液中的生物应用。
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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Silvia Cavagnero其他文献
Silvia Cavagnero的其他文献
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{{ truncateString('Silvia Cavagnero', 18)}}的其他基金
Development of a Laser-Assisted NMR Technology for the Atomic-Resolution Analysis of Medically Relevant Biomolecules in Solution at Submicromolar Concentration
开发激光辅助核磁共振技术,对亚微摩尔浓度溶液中医学相关生物分子进行原子分辨率分析
- 批准号:
10020189 - 财政年份:2018
- 资助金额:
$ 23.5万 - 项目类别:
Development of a Laser-Assisted NMR Technology for the Atomic-Resolution Analysis of Medically Relevant Biomolecules in Solution at Submicromolar Concentration
开发激光辅助核磁共振技术,对亚微摩尔浓度溶液中医学相关生物分子进行原子分辨率分析
- 批准号:
10242819 - 财政年份:2018
- 资助金额:
$ 23.5万 - 项目类别:
Development of LED-Assisted NMR Technologies for the Atomic-Resolution Analysis of Medically Relevant Biomolecules in Solution at Submicromolar Concentration
开发 LED 辅助 NMR 技术,对亚微摩尔浓度溶液中的医学相关生物分子进行原子分辨率分析
- 批准号:
10659378 - 财政年份:2018
- 资助金额:
$ 23.5万 - 项目类别:
Development of Laser-Mediated Hyper-Sensitive NMR in Liquids
激光介导液体超灵敏核磁共振的发展
- 批准号:
8757756 - 财政年份:2014
- 资助金额:
$ 23.5万 - 项目类别:
Development of Laser-Mediated Hyper-Sensitive NMR in Liquids
激光介导液体超灵敏核磁共振的发展
- 批准号:
8898152 - 财政年份:2014
- 资助金额:
$ 23.5万 - 项目类别:
Analysis of De Novo Protein Folding by Fluorescence Resonance Energy Transfer
通过荧光共振能量转移分析从头蛋白质折叠
- 批准号:
8373308 - 财政年份:2012
- 资助金额:
$ 23.5万 - 项目类别:
Analysis of De Novo Protein Folding by Fluorescence Resonance Energy Transfer
通过荧光共振能量转移分析从头蛋白质折叠
- 批准号:
8550099 - 财政年份:2012
- 资助金额:
$ 23.5万 - 项目类别:
Analysis of De Novo Protein Folding by Fluorescence Resonance Energy Transfer
通过荧光共振能量转移分析从头蛋白质折叠
- 批准号:
8852633 - 财政年份:2012
- 资助金额:
$ 23.5万 - 项目类别:
Analysis of De Novo Protein Folding by Fluorescence Resonance Energy Transfer
通过荧光共振能量转移分析从头蛋白质折叠
- 批准号:
8668100 - 财政年份:2012
- 资助金额:
$ 23.5万 - 项目类别:
CONFORMATION OF HSP70-BOUND PEPTIDE SUBSTRATES PROBED USING NMR SPECTROSCOPY
使用核磁共振波谱探测 HSP70 结合肽底物的构象
- 批准号:
8361245 - 财政年份:2011
- 资助金额:
$ 23.5万 - 项目类别:
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