Instrumentation Development: MS Array for Quantitative Proteomics

项目摘要/摘要 在分子水平上对人类疾病的研究因人类疾病技术的快速进步而受益匪浅 基因组测序。然而，有效利用当前和未来基因组信息的障碍 涉及与基因功能分配相关的挑战。蛋白质组学领域已经兴起 帮助解决这一需求，因为蛋白质是细胞中主要的功能分子。测量 蛋白质组内容，包括蛋白质表达水平、翻译后修饰和蛋白质相互作用 能够产生与基因功能相关的批判性见解。蛋白质组学研究中的主要挑战与 样品的复杂性和必须执行测量的宽动态范围。总体而言 就术语而言，这些需求比基因组学中遇到的要大得多，因为蛋白质丰度可以有很大的差异。 与基因相比，蛋白质在物理性质上有更广泛的多样性，蛋白质 没有信号放大的手段，也没有类似于基因的Watson-Crick碱基配对。因此，该领域 蛋白质组学的技术很大程度上基于多肽的质谱学测量，因为这些 测量显示了大规模蛋白质识别和定量的能力。然而，由于 每种蛋白质平均可以产生50-100个多肽，对多肽混合物的测量要多得多 比对蛋白质的测量复杂。此外，当前依赖数据的测量策略导致 显著压缩可实现的动态范围，因为此类MS/MS测量仅在正常情况下 对观察到的丰度较高的多肽是可行的。理想情况下，蛋白质组范围的消化中的每个肽都会 进行MS/MS以从蛋白质组学实验中获得最大信息。这项工程将会取得进展 通过开发能够高性能的质谱计阵列进行大规模蛋白质组学的能力 分辨率MS/MS采集比当前最先进的技术快一个数量级。多伦多 在该项目下开发的阵列技术将涉及离子回旋共振质谱学 这将产生更高的质量分辨率、更高的质量测量精度以及更高的吞吐量 收购。结果，在一个给定的实验中，可以识别一个数量级或更多的多肽 这将极大地增加每个蛋白质组分析的信息量以及动态 可以研究的蛋白质的范围。

PROJECT SUMMARY/ABSTRACT The study of human diseases at the molecular level has benefitted greatly by rapid advances in technology for genome sequencing. However, impediments for effective utilization of current and future genomic information relates to challenges associated with functional assignment of genes. The field of proteomics has arisen to help address this need, since proteins are the predominant functional molecules in cells. Measurement of proteomic content, including protein expression levels, posttranslational modifications and protein interactions can yield critical insight relevant to gene function. The major challenges in proteomics research relate to the complexity of samples and the wide dynamic range over which measurements must be performed. In general terms, these demands are far greater than encountered in genomics since, protein abundances can vary much more than genes do, proteins have much wider diversity in physical properties than genes do, and proteins have no means for signal amplification, nor analogous Watson-Crick base pairing as genes do. Thus, the field of proteomics employs technology largely based on mass spectrometry measurements of peptides since these measurements have shown capabilities for large-scale protein identification and quantitation. However, since each protein can produce on average, 50-100 peptides, measurements of peptide mixtures are far more complex than measurements on proteins. In addition, current data-dependent measurement strategies lead to significant compression of achievable dynamic range, since such MS/MS measurements are only normally feasible on peptides observed with higher abundance. Ideally, every peptide in a proteome-wide digest would be subjected to MS/MS to gain maximal information from proteomics experiments. This project will advance capabilities for large-scale proteomics through the development of a mass spectrometer array capable of high resolution MS/MS acquisition an order of magnitude faster than current state-of-the art technology. The MS array technology to be developed under this project will involve ion cyclotron resonance mass spectrometry that will yield higher mass resolving power, higher mass measurement accuracy as well as higher throughput acquisition. As a result, an order of magnitude or more peptides can be identified during a given experiment which will dramatically increase the information content of each proteomics analysis as well as the dynamic range of proteins that can be studied.