Purchase of a nanoLC-QTRAP Mass Spectrometer for Targeted Proteomics Studies

购买 nanoLC-QTRAP 质谱仪用于靶向蛋白质组学研究

• 批准号: 8447770
• 负责人: Sonja Hess
• 金额: $ 60万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8447770
• 关键词: Affect; Antibodies; Biological; Cell physiology; Complex; Coupled; Cullin Proteins; Embryo; Enzymes; Functional disorder; Funding; Gene Expression; Gene Proteins; Health; Histones; Human; Immunoblotting; Laboratories; Logic; Malignant Neoplasms; Mass Spectrum Analysis; Methods; Mitochondria; Molecular; Monitor; Mus; Mutate; Mutation; Nerve Degeneration; Neurologic; Orthologous Gene; Pathogenesis; Pathway interactions; Peptides; Proteins; Proteome; Proteomics; Research Personnel; Research Project Grants; Risk; Saccharomycetales; Sampling; Schizophrenia; Science; Sea Urchins; Secure; Synapses; System; T cell differentiation; Therapeutic; Woman; Yeasts; base; chemical genetics; genetic regulatory protein; human disease; immune activation; insight; instrument; mass spectrometer; mouse model; multiple reaction monitoring; offspring; response; transcription factor; ubiquitin ligase

DESCRIPTION (provided by applicant): This application is to secure funds to purchase an AB SCIEX QTRAP 5500 PRO triple quadrupole (QQQ) mass spectrometer coupled with an Eksigent nanoLC to carry out analyses based on multiple- reaction monitoring (MRM). MRM is an extremely sensitive, quantitative, and reproducible method for monitoring the levels of targeted, low abundance proteins in a complex biological sample. Conceptually it is similar to performing quantitative immunoblots but without the need to tag or raise antibodies against the targeted proteins. In the long term, this instrument will enable many research projects for which quantitative protein science is a core component. This application comprises eight specific projects. The Deshaies laboratory will quantify the levels of 64 proteins from the cullin-RING ubiquitin ligase (CRL) network in yeast bearing mutations in the Nedd8/Rub1 conjugating and deconjugating enzymes. This analysis will provide insights into the molecular logic of this key regulatory system which has been implicated in the pathogenesis of cancer. The Rothenberg laboratory will study a defined set of transcription factors in mice that will provide fundamental insight in T-cell differentiation. In addition, the Davidson laboratory will study a defined set of transcription factors in sea urchins that will delineate the spatial specifications of gene expression in sea urchin embryos. The Kennedy laboratory will use MRM to quantify synaptic regulatory proteins. The Hess lab will use mass spectrometry to quantify select acetylated histone peptides. The Chan laboratory will employ MRM to evaluate specific changes in the mitochondrial proteome in which cycles of mitochondrial fission and fusion or mitophagy are disrupted. The Patterson lab will study a set of 14 target proteins in an established mouse model of maternal immune activation, a condition known to increase the risk of schizophrenia in the offspring of affected women by 3-7 fold. Insights in the molecular pathways that trigger this pathophysiology will suggest new targets for cures. The Varshavsky lab will quantify a set of previously defined substrates of the N-end rule pathway in budding yeast using MRM. This will provide molecular insight into the neurological deficits that result when the orthologous gene is mutated in humans. The availability of a state-of-the-art QTRAP 5500 mass spectrometer will enable these Caltech researchers to employ targeted MRM to quantitatively monitor changes in proteomes in response to a variety of chemical, genetic, and environmental perturbations. All of the projects relate to critical subcellular processes and many of the genes and proteins to be studied are targets of human therapeutics or are causative agents in human diseases including neurodegeneration, schizophrenia, and cancer. Thus, the proposed studies have the potential to have a great impact on human health.

描述（由申请人提供）：本申请旨在获得资金，以购买与Eksigent nanoLC耦合的AB SCIEX QTRAP 5500 PRO三重四极杆（QQQ）质谱仪，以进行基于多反应监测（MRM）的分析。MRM是一种非常灵敏、定量和可重复的方法，用于监测复杂生物样品中靶向低丰度蛋白质的水平。从概念上讲，它类似于进行定量免疫印迹，但不需要标记或产生针对靶蛋白的抗体。从长远来看，该仪器将使许多以定量蛋白质科学为核心组成部分的研究项目成为可能。该应用程序包括八个具体项目。Deshaies实验室将对Nedd 8/Rub 1结合和去结合酶突变酵母中cullin-RING泛素连接酶（CRL）网络中的64种蛋白质水平进行定量。这项分析将提供深入了解这个关键的调控系统，这已牵连在癌症的发病机制的分子逻辑。Rothenberg实验室将在小鼠中研究一组确定的转录因子，这将为T细胞分化提供基本的见解。此外，戴维森实验室将研究一组定义的 这将描绘海胆胚胎中基因表达的空间规格。肯尼迪实验室将使用MRM来量化突触调节蛋白。赫斯实验室将使用质谱法来定量选择乙酰化组蛋白肽。Chan实验室将采用MRM来评估线粒体蛋白质组的特定变化，其中线粒体分裂和融合或线粒体自噬的周期被破坏。Patterson实验室将在一个已建立的母体免疫激活小鼠模型中研究一组14种靶蛋白，这种情况已知会使受影响女性的后代患精神分裂症的风险增加3-7倍。对触发这种病理生理学的分子途径的深入了解将为治疗提供新的靶点。Varshavsky实验室将使用MRM量化芽殖酵母中N端规则途径的一组先前定义的底物。这将提供分子洞察神经缺陷时，导致orthopathy基因突变的人类。最先进的QTRAP 5500质谱仪的可用性将使这些加州理工学院的研究人员能够采用有针对性的MRM来定量监测蛋白质组响应各种化学，遗传和环境扰动的变化。所有这些项目都涉及关键的亚细胞过程，许多待研究的基因和蛋白质都是人类治疗的目标，或者是人类疾病（包括神经变性，精神分裂症和癌症）的病原体。因此，拟议的研究有可能对人类健康产生重大影响。