Instrumentation for Quantitative Phosphoproteomics and Acetylomics

定量磷酸化蛋白质组学和乙酰组学仪器

• 批准号: 8636120
• 负责人: Martin Arthur Moseley
• 金额: $ 32万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-15 至 2015-05-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8636120
• 关键词: Area; Basic Science; Biological Markers; Biomedical Research; Blood capillaries; Clinical Sciences; Clinical Trials; Communicable Diseases; Complex; Computer software; Coupling; Data Analyses; Disease; Electrospray Ionization; Funding; Health; Informatics; Lead; Mass Spectrum Analysis; Metabolic Diseases; Neurology; Organism; Patients; Peptides; Performance; Pharmaceutical Preparations; Pharmacotherapy; Phenotype; Proteins; Proteomics; Research Infrastructure; Research Project Grants; Resolution; Sampling; Sickle Cell Anemia; System; Systems Analysis; Technology; Time; United States National Institutes of Health; Water; base; biological systems; capillary; improved; instrument; instrumentation; liquid chromatography mass spectrometry; mass spectrometer; medical schools; nanoscale; oncology; research study; response

DESCRIPTION (provided by applicant): Major biomedical discoveries have been made possible with advanced 'omic technologies, and continued technological advances in 'omic technologies are essential for a more complete physicochemical phenotyping (qualitative and quantitative characterizations) of normal and perturbed biological systems. Biological systems are overwhelming complex, and relatively subtle perturbations in the physicochemical status of these systems clearly lead to profound changes in the health of the organism. The Duke Proteomics Facility (DPCF) is tasked with supporting the protein/peptide characterization needs of the seven Basic Science Departments and the eleven Clinical Sciences Departments of the School of Medicine. The DPCF provides capabilities for mass spectrometry based proteomics for protein identification and protein quantitation, with the wide scope of capabilities required t support a variety of basic science studies and the significant scale of capacity to support clinica studies, specifically including biomarker discovery and biomarker verification experiments. The DPCF has ~ $5,000,000 in dedicated analytical and informatic hardware/software, and a staff of 7 (5 with PhDs, 6 of 7 full time staff). The DPCF has the pipeline technologies critical for LC/LC/MS/MS analysis of cellular proteins and peptides, including a robust and semi-automated analysis pipeline for proteomic, phosphoproteomic and acetylomics studies. In order to more completely physiochemically phenotype phosphoproteomes and acetylomes, the DPCF needs both additional state-of-the-art capabilities and increased capacity, both of which would be provided by the proposed electrospray ionization high resolution, accurate mass tandem mass spectrometer (Q-Exactive, Thermo Scientific). Analyses of identical samples in the DPCF and on a Q-Exactive show the requested instrument provides ~9X increase in coverage of the acetylome and ~ 7X increase in coverage of the phosphoproteome (at the PTM-modified peptide level). Coupling an existing multidimensional nanoscale capillary ultraperformance UPLC/UPLC system (Waters NanoAcquity with Multidimensional Technology) in the DPCF and the requested tandem mass spectrometer (Q-Exactive) will provide the absolute state-of-the-art in instrumentation for phosphoproteomic and acetylomics analyses. The research projects enabled by this proposal will advance our understanding of mechanisms of disease and drug therapy in areas including neurology, oncology, sickle cell disease, infectious diseases, and metabolic diseases. Ultimately, the instrumentation will improve clinical trials and patient therapies through the discovery and characterization of protein 'biomarkers' of health/disease/drug response. The requested system will provide unique performance capabilities that are synergistically aligned with the existing proteomic analysis systems and the comprehensive, dedicated informatics infrastructure in the DPCF, enabling Duke to leverage to the fullest extent the extensive expertise, robust data analysis capabilities, and plethora of important NIH- funded biomedical research opportunities at Duke.

描述（由申请人提供）：先进的“组学”技术使重大生物医学发现成为可能，而“组学”技术的持续技术进步对于正常和受干扰生物系统的更完整的物理化学表型（定性和定量表征）至关重要。生物系统是极其复杂的，这些系统的物理化学状态中相对微妙的扰动显然会导致有机体健康的深刻变化。杜克大学蛋白质组学设施（DPCF）的任务是支持医学院七个基础科学系和十一个临床科学系的蛋白质/肽表征需求。DPCF提供基于质谱的蛋白质组学功能，用于蛋白质鉴定和蛋白质定量，具有支持各种基础科学研究所需的广泛能力和支持临床研究的显著规模的能力，特别是包括生物标志物发现和生物标志物验证实验。DPCF拥有约500万美元的专用分析和信息硬件/软件，7名员工（5名拥有博士学位，7名全职员工中有6名）。DPCF拥有对细胞蛋白质和肽的LC/LC/MS/MS分析至关重要的流水线技术，包括用于蛋白质组学、磷酸化蛋白质组学和乙酰组学研究的强大的半自动分析流水线。为了更完整地进行磷酸化蛋白质组和乙酰化酶的物理化学表型，DPCF需要额外的最先进的能力和增加的容量，这两者都将由提议的电喷雾电离高分辨率、精确的质串联质谱仪（Q-Exactive, Thermo Scientific）提供。在DPCF和Q-Exactive中对相同样品的分析表明，所要求的仪器提供了~9X的乙酰酶组覆盖率和~ 7X的磷蛋白组覆盖率（在ptm修饰的肽水平上）。DPCF中现有的多维纳米级毛细管超高性能UPLC/UPLC系统（Waters NanoAcquity with multidimensional Technology）与所要求的串联质谱仪（Q-Exactive）相结合，将为磷蛋白质组学和乙酰组学分析提供绝对先进的仪器。该提案所支持的研究项目将促进我们对神经病学、肿瘤学、镰状细胞病、传染病和代谢性疾病等领域的疾病和药物治疗机制的理解。最终，该仪器将通过发现和表征健康/疾病/药物反应的蛋白质“生物标志物”来改善临床试验和患者治疗。所要求的系统将提供独特的性能能力，与现有的蛋白质组学分析系统和DPCF中全面、专用的信息学基础设施协同一致，使杜克大学能够充分利用杜克大学广泛的专业知识、强大的数据分析能力和大量重要的NIH资助的生物医学研究机会。

项目成果

The Aspergillus fumigatus septins play pleiotropic roles in septation, conidiation, and cell wall stress, but are dispensable for virulence.

烟曲霉脓毒症在分隔、分生孢子形成和细胞壁应激中发挥多效性作用，但对于毒力来说是可有可无的。

• DOI: 10.1016/j.fgb.2015.05.014
• 发表时间: 2015
• 期刊: Fungal genetics and biology : FG & B
• 作者: Vargas-Muñiz,JoséM;Renshaw,Hilary;Richards,AmberD;Lamoth,Frédéric;Soderblom,ErikJ;Moseley,MArthur;Juvvadi,PraveenR;Steinbach,WilliamJ
• 通讯作者: Steinbach,WilliamJ

Identification and mutational analyses of phosphorylation sites of the calcineurin-binding protein CbpA and the identification of domains required for calcineurin binding in Aspergillus fumigatus.

• DOI: 10.3389/fmicb.2015.00175
• 发表时间: 2015
• 期刊: Frontiers in microbiology
• 影响因子: 5.2
• 作者: Juvvadi PR;Ma Y;Richards AD;Soderblom EJ;Moseley MA;Lamoth F;Steinbach WJ
• 通讯作者: Steinbach WJ

Vagal nerve stimulation modifies neuronal activity and the proteome of excitatory synapses of amygdala/piriform cortex.

• DOI: 10.1111/jnc.13931
• 发表时间: 2017-03
• 期刊: Journal of neurochemistry
• 影响因子: 4.7
• 作者: Alexander GM;Huang YZ;Soderblom EJ;He XP;Moseley MA;McNamara JO
• 通讯作者: McNamara JO

Dephosphorylation of the Core Septin, AspB, in a Protein Phosphatase 2A-Dependent Manner Impacts Its Localization and Function in the Fungal Pathogen Aspergillus fumigatus.

• DOI: 10.3389/fmicb.2016.00997
• 发表时间: 2016
• 期刊: Frontiers in microbiology
• 影响因子: 5.2
• 作者: Vargas-Muñiz JM;Renshaw H;Richards AD;Waitt G;Soderblom EJ;Moseley MA;Asfaw Y;Juvvadi PR;Steinbach WJ
• 通讯作者: Steinbach WJ

The tail domain of the Aspergillus fumigatus class V myosin MyoE orchestrates septal localization and hyphal growth.

烟曲霉 V 类肌球蛋白 MyoE 的尾部结构域协调隔膜定位和菌丝生长。

• DOI: 10.1242/jcs.205955
• 发表时间: 2018
• 期刊: Journal of cell science
• 影响因子: 4
• 作者: Renshaw,Hilary;Vargas-Muñiz,JoséM;Juvvadi,PraveenR;Richards,AmberD;Waitt,Greg;Soderblom,ErikJ;Moseley,MArthur;Steinbach,WilliamJ
• 通讯作者: Steinbach,WilliamJ