Fluorescence lifetime-based single fluorophore biosensors of post-translational modification enzyme activity
基于荧光寿命的翻译后修饰酶活性单荧光团生物传感器
基本信息
- 批准号:9359690
- 负责人:
- 金额:$ 34.52万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2017
- 资助国家:美国
- 起止时间:2017-08-01 至 2020-07-31
- 项目状态:已结题
- 来源:
- 关键词:AcetyltransferaseAdvanced DevelopmentBehaviorBindingBiologicalBiological ModelsBiologyBiophysicsBiosensorBreast Cancer CellCalorimetryCell modelCell physiologyCellsComplexCultured CellsCysteineDataDevelopmentDiseaseEnergy TransferEnvironmentEnzyme Inhibitor DrugsEnzymesEventFamilyFluorescenceFunctional disorderGene ExpressionGoalsGoldGrantImageImageryImaging technologyIn VitroIndividualIntegrin Signaling PathwayLabelLeadLibrariesLightLiteratureLocationMalignant NeoplasmsMapsMeasuresMethodsMicroscopyModificationNaturePeptidesPhosphopeptidesPhosphorylated PeptidePhosphorylationPhosphotransferasesPopulationPost Translational Modification AnalysisPost-Translational Protein ProcessingProcessProtein KinaseProtein Tyrosine KinaseProteinsProteomicsReactionReaderRecombinantsReportingResearchResolutionRoleScanningSignal PathwaySignal TransductionSpectrum AnalysisStimulusSurface Plasmon ResonanceSystemTechniquesTechnologyTestingTherapeuticTimeTitrationsUnited States National Institutes of HealthValidationWorkbasecell behaviorcellular imagingdesignemergency service responderenzyme activityenzyme substrateexperienceexperimental studyfluorescence lifetime imagingfluorophoreimaging modalityimaging probeinnovationinsightinstrumentationnovelnovel strategiesprogramsprotein protein interactionresponsesingle cell analysisspatiotemporalsrc Homology Region 2 Domaintooluptakevirtual
项目摘要
Despite decades of research on post-translational modification enzymes, little is known about their dynamic,
spatiotemporal activities on a subcellular level—and yet the nature of these subcellular activities is often key to
understanding their dysfunction in cancer. The goal of this project is to develop a fluorescence lifetime-based
technology to map those activities in living cells at subcellular resolution. This approach is based on robust
preliminary data in which we demonstrate feasibility for detecting tyrosine kinase activity (an important PTM
in many cancers) via highly resolved fluorescence lifetime shifts for three peptide probes (for Abl, Src-family,
and Syk kinases), including multiplexed analysis of two probes at a time. In Aim 1, we will fully characterize the
physiochemical mechanism that gives rise to the shift, which involves binding of the phosphorylated peptide
product of the kinase reaction to an SH2 domain. This will allow us to understand and predict lifetime shift
effects, enabling targeted probe design. In Aim 2, we will characterize and further develop the multiplexed
analysis of multiple probes, using a biologically relevant model system (the integrin signaling pathway) to
validate the utility of the method. In Aim 3, we will broaden the scope of the technology to detect other PTM
enzyme activities, starting with Ser/Thr kinases and acetyltransferases. The work described in this proposal
will deliver 1) a suite of novel, complementary probes for PTM enzyme function in cells, 2) a strategy to develop
additional probes, 3) high-throughput in vitro methods for detecting PTMs via fluorescence lifetime shifts, and
4) cutting edge methods for imaging multiple probes in the same cell. These tools will make unique
contributions to the understanding of PTM dynamics in the cell and to the elucidation of roles for subcellular
kinase signaling in disease.
尽管对翻译后修饰酶进行了数十年的研究,但对其动态,
然而,这些亚细胞活动的性质往往是关键,
了解他们在癌症中的功能障碍。该项目的目标是开发一种基于荧光寿命的
这项技术可以在亚细胞分辨率下绘制活细胞中的这些活动。该方法基于鲁棒性
初步数据,我们证明了检测酪氨酸激酶活性的可行性(一个重要的PTM
在许多癌症中)通过三种肽探针(Abl,Src-家族,
和Syk激酶),包括一次两个探针的多重分析。在目标1中,我们将充分描述
引起转变的生理化学机制,涉及磷酸化肽的结合
激酶与SH 2结构域反应的产物。这将使我们能够理解和预测寿命的变化
影响,使有针对性的探针设计。在目标2中,我们将表征并进一步开发多路复用
多个探针的分析,使用生物学相关的模型系统(整合素信号通路),
验证了该方法的实用性。在目的3中,我们会扩大技术的范围,以侦测其他PTM
酶活性,从Ser/Thr激酶和乙酰转移酶开始。本提案中所述的工作
将提供1)一套新的,互补的探针PTM酶功能的细胞,2)战略,
另外的探针,3)通过荧光寿命变化检测PTM的高通量体外方法,和
4)在同一细胞中成像多个探针的尖端方法。这些工具将使独特的
有助于理解PTM在细胞中的动力学和阐明亚细胞的作用,
疾病中的激酶信号传导。
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)
数据更新时间:{{ journalArticles.updateTime }}
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
数据更新时间:{{ journalArticles.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ monograph.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ sciAawards.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ conferencePapers.updateTime }}
{{ item.title }}
- 作者:
{{ item.author }}
数据更新时间:{{ patent.updateTime }}
Laurie L. Parker其他文献
Laurie L. Parker的其他文献
{{
item.title }}
{{ item.translation_title }}
- DOI:
{{ item.doi }} - 发表时间:
{{ item.publish_year }} - 期刊:
- 影响因子:{{ item.factor }}
- 作者:
{{ item.authors }} - 通讯作者:
{{ item.author }}
{{ truncateString('Laurie L. Parker', 18)}}的其他基金
Multiplexed proteomics-based kinase assay development
基于多重蛋白质组学的激酶测定开发
- 批准号:
10810004 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
Multiplexed proteomics-based kinase assay development
基于多重蛋白质组学的激酶测定开发
- 批准号:
10467462 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
Multiplexed proteomics-based kinase assay development
基于多重蛋白质组学的激酶测定开发
- 批准号:
10615893 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
Multiplexed proteomics-based kinase assay development
基于多重蛋白质组学的激酶测定开发
- 批准号:
10793244 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
High-Throughput Screening Platform for Cancer Drug Discovery
癌症药物发现的高通量筛选平台
- 批准号:
10432025 - 财政年份:2020
- 资助金额:
$ 34.52万 - 项目类别:
High-Throughput Screening Platform for Cancer Drug Discovery
癌症药物发现的高通量筛选平台
- 批准号:
10163816 - 财政年份:2020
- 资助金额:
$ 34.52万 - 项目类别:
Biosensor Assay to Screen for Signaling Pathway Inhibition in Cancer
用于筛选癌症信号通路抑制的生物传感器测定
- 批准号:
9445128 - 财政年份:2017
- 资助金额:
$ 34.52万 - 项目类别:
Biosensor Assay to Screen for Signaling Pathway Inhibition in Cancer
用于筛选癌症信号通路抑制的生物传感器测定
- 批准号:
8956420 - 财政年份:2014
- 资助金额:
$ 34.52万 - 项目类别:
Biosensor Assay to Screen for Signaling Pathway Inhibition in Cancer
用于筛选癌症信号通路抑制的生物传感器测定
- 批准号:
9076374 - 财政年份:2014
- 资助金额:
$ 34.52万 - 项目类别:
Multiplexed Kinase Biosensor Technology to Detect Leukemia Signaling with Mass Sp
多重激酶生物传感器技术通过 Mass Sp 检测白血病信号传导
- 批准号:
8930087 - 财政年份:2014
- 资助金额:
$ 34.52万 - 项目类别:
相似海外基金
ADVANCED DEVELOPMENT OF LQ A LIPOSOME-BASED SAPONIN-CONTAINING ADJUVANT FOR USE IN PANSARBECOVIRUS VACCINES
用于 Pansarbecovirus 疫苗的 LQ A 脂质体含皂苷佐剂的先进开发
- 批准号:
10935820 - 财政年份:2023
- 资助金额:
$ 34.52万 - 项目类别:
ADVANCED DEVELOPMENT OF BBT-059 AS A RADIATION MEDICAL COUNTERMEASURE FOR DOSING UP TO 48H POST EXPOSURE"
BBT-059 的先进开发,作为辐射医学对策,可在暴露后 48 小时内进行给药”
- 批准号:
10932514 - 财政年份:2023
- 资助金额:
$ 34.52万 - 项目类别:
Advanced Development of a Combined Shigella-ETEC Vaccine
志贺氏菌-ETEC 联合疫苗的先进开发
- 批准号:
10704845 - 财政年份:2023
- 资助金额:
$ 34.52万 - 项目类别:
Advanced development of composite gene delivery and CAR engineering systems
复合基因递送和CAR工程系统的先进开发
- 批准号:
10709085 - 财政年份:2023
- 资助金额:
$ 34.52万 - 项目类别:
Advanced development and validation of an in vitro platform to phenotype brain metastatic tumor cells using artificial intelligence
使用人工智能对脑转移肿瘤细胞进行表型分析的体外平台的高级开发和验证
- 批准号:
10409385 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
ADVANCED DEVELOPMENT OF A VACCINE FOR PANDEMIC AND PRE-EMERGENT CORONAVIRUSES
针对大流行和突发冠状病毒的疫苗的高级开发
- 批准号:
10710595 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
Advanced development and validation of an in vitro platform to phenotype brain metastatic tumor cells using artificial intelligence
使用人工智能对脑转移肿瘤细胞进行表型分析的体外平台的高级开发和验证
- 批准号:
10630975 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
ADVANCED DEVELOPMENT OF A VACCINE CANDIDATE FOR STAPHYLOCOCCUS AUREUS INFECTION
金黄色葡萄球菌感染候选疫苗的高级开发
- 批准号:
10710588 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别:
ADVANCED DEVELOPMENT OF A VACCINE FOR PANDEMIC AND PRE-EMERGENT CORONAVIRUSES
针对大流行和突发冠状病毒的疫苗的高级开发
- 批准号:
10788051 - 财政年份:2022
- 资助金额:
$ 34.52万 - 项目类别: