Establishment of a High Performance UPLC-QSTAR Elite Proteomics Platform at Unive

大学高性能UPLC-QSTAR Elite蛋白质组学平台的建立

• 批准号: 7588397
• 负责人: Sixue Chen
• 金额: $ 32.6万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7588397
• 关键词: Arts; Biological Markers; Biomedical Research; Biotechnology; Brain Injuries; Communicable Diseases; Complement; Complex; Complex Mixtures; Computer software; Data Analyses; Development; Dimensions; Disease; Experimental Designs; Florida; Functional disorder; Funding; Gel; Generations; Grant; Health; Human; Institution; Ions; Isotope Labeling; Liquid Chromatography; Malignant Neoplasms; Mass Spectrum Analysis; Neurodegenerative Disorders; Peptides; Performance; Phosphorylation; Post-Translational Protein Processing; Productivity; Protein Array; Proteins; Proteome; Proteomics; Public Health; Reproducibility; Research; Research Personnel; Resolution; Sampling; Services; Shotguns; Speed; Staging; System; Technology; Time; United States National Institutes of Health; Universities; Water; base; disorder prevention; drug development; environmental toxicology; human disease; improved; instrument; instrumentation; mass spectrometer; new technology; public health relevance; research study; transmission process; two-dimensional

DESCRIPTION (provided by applicant): This shared instrumentation application seeks funding for the establishment of a high performance modern proteomics platform to enhance biomedical research at the University of Florida (UF). The platform consists of an Applied Biosystems new generation QSTAR Elite quadrupole time-of-flight (QTOF) mass spectrometer and associated nanoflow two-dimensional ultra performance liquid chromatography (UPLC) (manufactured by Waters Inc.) and ProteinPilot 2.0 software package for data analysis. We propose to operate this integrated nanoflow UPLC-QTOF MS/MS system for separation and analysis of complex peptide (and protein) mixtures as part of a large group of over 20 NIH funded grants, from which eight representative groups were selected to demonstrate the pressing needs for the state-of-the-art proteomics technology. The most critical features of this new system are: significant improvement in front-end separation and the number of mass spectra that can be acquired, overall sensitivity, resolution and reproducibility, dynamic range, characterization of low abundance peptides, smart collision energetics, ion transmission/accumulation, and quantitation software features. The unique capabilities of this proteomics platform will be directed at a large array of protein and proteomic applications where LC-MS is used for protein/peptide identification, accurate quantitation of complex mixtures, as well as the determination of biologically significant posttranslational modifications, such as protein phosphorylation. Because of the limitations of 2D gel based proteomics, recent years have seen increasing demands for shotgun proteomics implementing multiple dimension LC-MS with or without isotope labeling. Such technology has been proven effective in expanding the proteome coverage and identifying low abundance proteins. However, it requires significant instrument time on a per experiment basis. Access to the existing LC-MS instrument in our Proteomics facility is limited due to the growing demands, and newer technologies outperform existing instruments in the Proteomics Division at the Interdisciplinary Center for Biotechnology Research (ICBR), the major research service facility on the UF campus. Therefore, we propose to establish this new technical platform to provide greater speed and performance for iTRAQ experiments and shotgun proteomics while complementing existing technologies in the facility and improving the sample throughput, experimental design, and productivity of the NIH projects that are examining key issues closely related to human health, including neurodegenerative disease, brain injury, cancer, biomarker discovery environmental toxicology, and infectious diseases. PUBLIC HEALTH RELEVANCE: Many human diseases and disorders are directly or indirectly caused by protein dysfunction. Protein biomarkers, many of which are present in low amounts and in a complex mixture of molecules, are indicative of disease states and/or are responsible for disease development. Acquisition of a high performance proteomics platform will greatly facilitate the NIH funded researchers at the University of Florida and the vinicity institutions to detect, identify and quantify the protein biomarkers at an early stage of disease, inform effective approaches to disease prevention, treatment and drug development, and thus greatly improve public health.

描述(由申请人提供)：该共享仪器申请寻求资金，以建立一个高性能的现代蛋白质组学平台，以加强佛罗里达大学(UF)的生物医学研究。该平台包括应用生物系统公司的新一代QSTAR Elite四极杆飞行时间(QTOF)质谱仪和相关的纳流二维超高效液相色谱(UPLC)(由Waters Inc.制造)。和ProteinPilot 2.0软件进行数据分析。我们建议使用这个集成的纳米流UPLC-QTOF MS/MS系统来分离和分析复杂的多肽(和蛋白质)混合物，作为NIH资助的20多个大小组的一部分，从这些资助中选择了8个具有代表性的小组来展示对最先进的蛋白质组学技术的迫切需求。这个新系统最关键的特点是：在前端分离和可获得的质谱数方面有显著改进，总体灵敏度、分辨率和重现性、动态范围、低丰度多肽的表征、智能碰撞能量学、离子传输/积累和定量软件功能。该蛋白质组学平台的独特能力将针对大量的蛋白质和蛋白质组应用，其中LC-MS用于蛋白质/肽的鉴定、复杂混合物的准确定量以及确定具有生物学意义的翻译后修饰，如蛋白质磷酸化。由于基于二维凝胶的蛋白质组学的局限性，近年来，人们对实现多维LC-MS的鸟枪式蛋白质组学的需求越来越大，无论是否有同位素标记。这种技术已经被证明在扩大蛋白质组覆盖范围和识别低丰度蛋白质方面是有效的。然而，在每次实验的基础上，它需要相当多的仪器时间。由于不断增长的需求，我们蛋白质组学设施中现有的LC-MS仪器的使用受到限制，而新技术的表现优于UF园区主要研究服务设施--跨学科生物技术研究中心(ICBR)蛋白质组学部门的现有仪器。因此，我们建议建立这个新的技术平台，为iTRAQ实验和猎枪蛋白质组学提供更快的速度和性能，同时补充设施中的现有技术，并提高NIH项目的样品吞吐量、实验设计和生产率，这些项目正在研究与人类健康密切相关的关键问题，包括神经退行性疾病、脑损伤、癌症、生物标记物发现、环境毒理学和传染病。公共卫生相关性：许多人类疾病和障碍是由蛋白质功能障碍直接或间接引起的。蛋白质生物标志物，其中许多存在于低含量和复杂的分子混合物中，指示疾病状态和/或负责疾病的发展。收购一个高性能的蛋白质组学平台将极大地方便美国国立卫生研究院资助的佛罗里达大学和葡萄酒研究所的研究人员在疾病的早期阶段检测、识别和量化蛋白质生物标志物，为疾病预防、治疗和药物开发提供有效的方法，从而极大地改善公众健康。

项目成果

Thiol-based redox proteins in abscisic acid and methyl jasmonate signaling in Brassica napus guard cells.

• DOI: 10.1111/tpj.12490
• 发表时间: 2014-05
• 期刊: The Plant journal : for cell and molecular biology
• 作者: Zhu M;Zhu N;Song WY;Harmon AC;Assmann SM;Chen S
• 通讯作者: Chen S

Quantitative proteomics of tomato defense against Pseudomonas syringae infection.

• DOI: 10.1002/pmic.201200402
• 发表时间: 2013-06
• 期刊: PROTEOMICS
• 影响因子: 3.4
• 作者: Parker, Jennifer;Koh, Jin;Yoo, Mi-Jeong;Zhu, Ning;Feole, Michelle;Yi, Sarah;Chen, Sixue
• 通讯作者: Chen, Sixue

Analysis of abscisic acid responsive proteins in Brassica napus guard cells by multiplexed isobaric tagging.

• DOI: 10.1016/j.jprot.2009.11.002
• 发表时间: 2010-02
• 期刊: Journal of proteomics
• 影响因子: 3.3
• 作者: Mengmeng Zhu;B. Simons;Ning Zhu;D. Oppenheimer;Sixue Chen