5500 Q-Trap Mass Spectrometer

描述（由申请人提供）：这是ABI-Sciex 5500 Qtrap的申请，用于支持8名NIH调查员的研究，他们正在进行生物途径，蛋白质复合物，蛋白质翻译后修饰和小生物活性肽在生命系统中的研究。与现有的4000 Qtrap相比，5500 Qtrap具有显著的灵敏度优势，其多反应离子监测（MRM）模式的灵敏度提高了5倍，MSMS分析的灵敏度提高了30倍。高灵敏度加上非常快的扫描速度（高达20000米/秒）是一个重大的进步。它允许监测100个MRM通道，并能够在每个MRM通道的峰值通过期间捕获确认的MSMS频谱。有了洗脱时间调度，在单次LC运行中监测的通道数量可以大大增加。该仪器非常适合定向蛋白质组学，其中感兴趣的蛋白质是明确定义的，无论是通过其他蛋白质质谱实验，从DNA微阵列分析，基因测序，或从其他生物学结果。在这种方法中，代表蛋白质的肽是根据长度（8-18个残基）、氨基酸组成（省略易氧化残基的肽- Cys、His、Met、Trp和Tyr）和序列独特性来仔细选择的。这些肽也可以通过制备稳定的同位素标记形式进行定量测量。这允许在单次LC运行中分析蛋白质/酶或蛋白质复合物的整个途径，从而了解蛋白质周转的作用。这些包括活化中性粒细胞中的蛋白酶体复合物，糖酵解和产生能量的线粒体途径中的酶，以及衰老过程中的晶状体结晶蛋白。大量的MRM通道也允许研究严重修饰的蛋白质，如囊性纤维化跨膜调节剂（我们已经观察到超过60个翻译后修饰）和晶态结晶蛋白。5500 Qtrap的高灵敏度还将允许测量小肽，如慢性阻塞性肺疾病中由肺部胶原蛋白产生的小肽，以便在更方便/可获得的液体（如血浆或尿液）中进行测量。所有这些实验的关键问题是，研究人员希望在正常丰度水平下对生物系统中的蛋白质进行分析。5500 Qtrap灵敏度的提高显著提高了这一能力。

DESCRIPTION (provided by applicant): This is an application for an ABI-Sciex 5500 Qtrap to support research by 8 named NIH investigators who are carrying out research on biological pathways, protein complexes, protein post-translational modifications and small biologically active peptides in living systems. The 5500 Qtrap has significant sensitivity advantages over the existing 4000 Qtrap with a 5-fold increase in the multiple reaction ion monitoring (MRM) mode and >30 fold for MSMS analysis. The high sensitivity plus the very fast scanning speed (up to 20,000 m/z per sec) is a significant advance. It allows the monitoring of 100 MRM channels with the ability to capture a confirmatory MSMS spectrum during the passage of a peak in each MRM channel. With elution time scheduling, the number of channels monitored in a single LC run can be substantially increased. This instrument is well suited to directed proteomics in which the proteins of interest are well-defined, either by other protein mass spectrometry experiments, from DNA microarray analysis, gene sequencing, or from other biological results. In this approach, peptides representing proteins are carefully selected based on length (8-18 residues), amino acid composition (omitting peptides with easily oxidized residues - Cys, His, Met, Trp and Tyr) and sequence uniqueness to act as surrogates. These peptides can also be measured quantitatively by the preparation of stable isotope-labeled forms. This allows whole pathways of proteins/enzymes or protein complexes to be analyzed in a single LC run and hence to understand the role of protein turnover. These include the proteasome complex in activated neutrophils, the enzymes in glycolysis and energy generating mitochondrial pathways, and the lens crystallins in aging. The large number of MRM channels also allows the study of heavily modified proteins such as the cystic fibrosis transmembrane regulator (we have observed over 60 post- translational modifications) and lens crystallins. The high sensitivity of the 5500 Qtrap will also allow measurement of small peptides such as those produced from collagen in the lung in chronic obstructive pulmonary disease to be measured in more convenient/accessible fluids such plasma or urine. The key issue in all these experiments is that the investigators want to carry out analyses on proteins in biological systems at normal abundance levels. The increased sensitivity of the 5500 Qtrap significantly improves that capability. PUBLIC HEALTH RELEVANCE: The requested instrument is much more sensitive than its predecessors and will allow investigators to examine the complexity of protein chemistry in living tissues. It will be applied to the study of mechanisms of neutrophilassociated damage in the lung, mitochondrial-associated damage in models of oxidative stress (atherosclerosis, diabetes and cancer), lens cataract disease in aging, pathway robustness in the light of gene damage/loss, and in Cystic Fibrosis. It's very well suited to obtaining quantitative data to better understand the role of the protein(s) in specific areas of public health.