Fluorous Quantitative Proteomics

荧光定量蛋白质组学

• 批准号: 7217109
• 负责人: Marvin S Yu
• 金额: $ 18.01万
• 依托单位: FLUOROUS TECHNOLOGIES, INC.
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-30 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7217109
• 关键词: analog; aniline dye; biotechnology; chemical synthesis; electrospray ionization mass spectrometry; fluorescent dye /probe; liquid chromatography; mass spectrometry; method development; oxidation reduction reaction; posttranslational modifications; protein quantitation /detection; proteomics; reagent /indicator; stable isotope

DESCRIPTION (provided by applicant): Project Summary/Abstract: The long-term goal of the project is to develop and then commercialize stable isotope reagents for quantitative proteomics using fluorous enrichment techniques. Successful demonstration in proteomics will be followed by extension into metabolomics and glycomics using a similar tagging strategy. Fluorous Technologies, Inc. (FTI) as the small business concern, and The Genomics Institute of the Novartis Research Foundation (GNF), as a collaborator, will synthesize fluorous isotope reagent tags (FLIRTs), develop protocols, benchmark vs. existing techniques, and demonstrate the FLIRT approach in the quantification of protein function. The specific aims of the grant are: 1) Synthesis of aniline based FLIRTs. Synthetic routes to 5 fluorous probes for chemical labeling of peptides and proteins will be synthesized based on a fluorous aniline core. A goal of 500 mg of each unlabeled compound and 150 mg of each labeled compound is targeted. 2) Synthesis of stable isotope labeled versions of the existing fluorous reagents. FLIRTs with the stable isotopes directly in the fluorous chain will be synthesized. The goal will be to obtain 100 mg of each compound. These compounds will be the direct stable isotope labeled analogs of fluorous peptide labeling reagents which FTI has previously produced. 3) Development of quantitative tagging protocols using the fluorous isotope labeled reagents and benchmarking vs. cICAT. Protocols for tagging and enrichment using FLIRTs will be developed using LC/MS with ESI detection. Quantification by FLIRT will then be benchmarked against cICAT. Performance measures will include enrichment efficiency, selectivity biases, selectivity enrichment, recovery of enriched species, and accuracy of quantitation. 4) Fluorous isotope reagent tagging (FLIRT) for relative quantification of a targeted subproteome. FLIRT technology will be applied to the quantification of specific classes of proteins within a complex sample, including the characterization of the redox state of the various cysteine residues within a protein. Successful enrichment and quantitation through a single label will demonstrate the flexibility of FLIRTs in analyzing targeted classes of proteins. FLIRTs will provide a novel method for quantitative proteomics which can be applied to numerous research areas including target discovery, biomarker discovery, and diagnostics development for various diseases affecting the human condition. Project Narrative: The goal of the project is to develop and commercialize reagents for quantitative proteomics. The successful commercialization of these products will enable researchers to elucidate biological pathways leading to new biomarkers and targets by which to develop novel diagnostics and therapeutics for various diseases that affect the human condition.

描述(由申请人提供)：项目摘要/摘要：该项目的长期目标是开发稳定的同位素试剂，然后利用氟浓缩技术将其商业化，用于定量蛋白质组学。在蛋白质组学的成功演示之后，将使用类似的标记策略扩展到代谢组学和糖组学。作为小型企业的氟技术公司(FTI)和诺华研究基金会(GNF)的基因组研究所将合成氟同位素试剂标签(FLORTT)，开发方案，对现有技术进行基准测试，并展示FLORTT方法在蛋白质功能量化中的应用。该基金的具体目标是：1)合成以苯胺为基础的调情对象。合成路线为5个用于多肽和蛋白质化学标记的含氟探针，将以氟苯胺为核心合成。目标是每种未标记化合物500毫克和每种标记化合物150毫克。2)现有氟试剂的稳定同位素标记物的合成。直接与氟链中的稳定同位素调情的将被合成。目标是获得每种化合物100毫克的含量。这些化合物将是FTI以前生产的氟肽标记试剂的直接稳定同位素标记类似物。3)开发使用氟同位素标记试剂的定量标记方案，并与cICAT进行基准比较。将使用带有ESI检测的LC/MS开发利用调情者进行标记和浓缩的协议。然后，将以CICAT为基准进行调情量化。绩效指标将包括浓缩效率、选择性偏差、选择性浓缩、浓缩物种的回收率和定量的准确性。4)用于目标亚蛋白质组相对定量的氟同位素试剂标记(FLORTT)。FLORTT技术将用于对复杂样品中特定类别的蛋白质进行量化，包括表征蛋白质中各种半胱氨酸残基的氧化还原状态。通过单一标记成功的浓缩和定量将证明调情者在分析目标类别的蛋白质时的灵活性。Flirts将为定量蛋白质组学提供一种新的方法，可应用于许多研究领域，包括靶点发现、生物标记物发现和各种影响人类状况的疾病的诊断开发。项目简介：该项目的目标是开发定量蛋白质组学试剂并将其商业化。这些产品的成功商业化将使研究人员能够阐明导致新的生物标记物和靶点的生物路径，从而为影响人类状况的各种疾病开发新的诊断和治疗方法。