Top-Down Proteomics: Ready for Prime Time?
Top-Down Proteomics: Ready for Prime Time?
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DOI：
10.1021/acs.analchem.7b04747
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发表时间：
2018-01-02
影响因子：
7.4
通讯作者：
Ge Y
中科院分区：
文献类型：
--
作者：
Chen B;Brown KA;Lin Z;Ge Y
除了遗传密码外，人类蛋白质组还因遗传变异、RNA转录物的选择性剪接和翻译后修饰（PTM）而变得多样化。6，7 2013年，术语“蛋白质型”被指定用于描述“单个基因的蛋白质产物可以被发现的所有不同的分子形式”，6消除了命名中的混乱，并加入了研究工作，以开发蛋白质型表征的方法。自上而下的蛋白质组学，分析完整的蛋白质没有消化，已被证明是一个首要的质谱（MS）技术的全球和全面的分析蛋白质。4，8，9自上而下的方法保留了完整的蛋白质质量信息，提供了蛋白质组的“鸟瞰”视图，并允许识别新的蛋白质型，深入序列表征和定量疾病相关的PTM。4，8，9尽管仍存在一些技术挑战，但过去5年的发展已将自上而下的蛋白质组学从主要针对性方法扩展到能够在多个样品中进行蛋白质组分析的方法。9现在，成千上万的蛋白质组可以使用高通量自上而下的蛋白质组学方法进行鉴定，表征和定量。10，11此外，仪器和片段化的发展使自上而下的蛋白质组学走上了未来发展的快车道。12随着商业高分辨率质谱仪的成功开发，例如solariX XR傅里叶变换离子回旋共振（FTICR）（布鲁克）、13 Orbitraps（赛默）和四极飞行时间（Q-TOF）（布鲁克和沃茨），这些仪器现已广泛应用于学术和工业实验室，用于自上而下的蛋白质组学。此外，太平洋西北国家实验室和国家高磁场实验室的21 T FTICR质谱仪配备了最先进的裂解方法，表现出前所未有的分辨率，采集速率和灵活的串联MS（MS/MS）功能，为自上而下的蛋白质组学从业者提供了巨大的潜力，以探索复杂的蛋白质组学应用。14，15反过来，从开发大规模和高通量工作流程中获得的方法和技术收益进一步增强了有针对性的分析，自上而下的蛋白质组学由此生根并蓬勃发展。因此，对自上而下的MS的兴趣大大增加，许多研究已经强调了自上而下的蛋白质组学在揭示疾病机制和发现新的生物标志物方面的潜力。4，8，16，17最近的一些评论已经概述了自上而下的蛋白质组学的技术要求，并描述了该领域的历史和基本原理及其在生物医学研究中的应用。4，8，9，18 - 20特别强调过去两年（2015 - 2017年）的出版物，本文综述了完整蛋白质样品制备，分离，MS/MS，数据采集策略，数据分析，天然MS和定量领域的最新技术趋势和发展。我们还强调了全球和有针对性的自上而下的蛋白质组学应用程序的最新应用，并得出结论，在该领域的前景。
Beyond the genetic code, the human proteome is greatly diversified by genetic variations, alternative splicing of RNA transcripts, and post-translational modifications (PTMs). 6, 7 In 2013, the term “proteoform” was designated to describe “all of the different molecular forms in which the protein product of a single gene can be found”, 6 clearing up the confusion in nomenclature and joining research efforts to develop methodologies for proteoform characterization. Top-down proteomics, which analyzes intact proteins without digestion, has proved to be a premier mass spectrometry (MS) technology for global and comprehensive analysis of proteoforms. 4, 8, 9 The top-down approach retains intact protein mass information, providing a “bird’s-eye” view of the proteome and allowing for identification of novel proteoforms, in-depth sequence characterization, and quantitation of disease-associated PTMs. 4, 8, 9 Although some technical challenges remain, development over the past 5 years has expanded top-down proteomics from a mostly targeted approach to one capable of proteoform-profiling across multiple samples. 9 Now, thousands of proteoforms can be identified, characterized, and quantified using high-throughput top-down proteomics approaches. 10, 11 Moreover, developments in instrumentation and fragmentation have positioned top-down proteomics in the fast lane for future progression. 12 With the successful development of commercial high-resolution mass spectrometers such as the solariX XR Fourier transform ion cyclotron resonance (FTICR)(Bruker), 13 Orbitraps (Thermo), and quadrupole time-of-flight (Q-TOFs)(Bruker and Waters), these instruments are now widely available in academic and industrial laboratories for top-down proteomics. Moreover, the 21 T FTICR mass spectrometers equipped with state-of-the-art fragmentation methods at the Pacific Northwest National Laboratory and National High Magnetic Field Laboratory have demonstrated unprecedented resolving power, acquisition rate, and flexible tandem MS (MS/MS) capabilities, providing enormous potential for top-down proteomics practitioners to probe complicated proteomics applications. 14, 15 Reciprocally, the methodological and technological gains from developing large-scale and high-throughput workflow have further empowered targeted analysis, from which top-down proteomics roots and thrives. As a result, interest in top-down MS has grown considerably and many studies have already underscored the potential of top-down proteomics for unraveling disease mechanisms and discovering novel biomarkers. 4, 8, 16, 17 A number of recent reviews have already given an overview of the technical requirements for top-down proteomics and delineated the history and fundamentals of the field as well as its application to biomedical research. 4, 8, 9, 18− 20 With a special emphasis on publications in the past 2 years (2015− 2017), this review examines recent technological trends and developments in the areas of intact protein sample preparation, separation, MS/MS, data acquisition strategies, data analysis, native MS, and quantitation. We also highlight recent applications for global and targeted top-down proteomics applications and conclude with outlooks in the field.
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影响因子：
7.4
作者：
Brodbelt JS
通讯作者：
Brodbelt JS
影响因子：
15
作者：
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影响因子：
56.9
作者：
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通讯作者：
Dumenil, Guillaume
影响因子：
7
作者：
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通讯作者：
Hunt, Donald F.