Analytical strategies for shotgun phosphoproteomics: Status and prospects |
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| Authors: | Haruna Imamura Masaki Wakabayashi Yasushi Ishihama |
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| Institution: | 1. State Key Laboratory of Virology,College of Life Sciences, Wuhan University, Wuhan, P.R. China;2. Key Laboratory of Analytical Chemistry for Biology and Medicine (Ministry of Education), Wuhan University, Wuhan, P.R. China;1. Division of Analytical Chemistry, Institute of Chemical Sciences, Bahauddin Zakariya University, Multan 60800, Pakistan;2. Institute of Analytical Chemistry and Radiochemistry, Leopold-Franzens University, Innrain 80-82, A-6020 Innsbruck, Austria;1. Department of Plant Nutrition, College of Resources and Environmental Sciences, China Agricultural University, Beijing, PR China;2. Department of Environmental Sciences & Engineering, College of Resources and Environmental Sciences, China Agricultural University, Beijing, PR China;1. Key Laboratory of Synthetic and Natural Function Molecule Chemistry of Ministry of Education, College of Chemistry and Materials Science, Northwest University, Xi''an, 710069, China;2. CAS Key Laboratory of Separation Science for Analytical Chemistry, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian, 116023, China;3. University of Chinese Academy of Sciences, Beijing, 100049, China;1. Liaoning Provincial Key Laboratory for Proteomics, Dalian Medical University, Dalian 116044, China;2. Key Laboratory of Separation Sciences for Analytical Chemistry, National Chromatographic R. & A. Center, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, Dalian 116023, China |
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| Abstract: | New analytical strategies for phosphoproteomics, both experimental and computational, have been rapidly introduced in recent years, leading to novel biological findings on the role of protein phosphorylation, which have in turn stimulated further development of the analytical techniques. In this review, we describe the development of analytical strategies for LC–MS/MS-based phosphoproteomics, focusing particularly on recent progress in phosphopeptide enrichment, LC–MS/MS measurement and the subsequent computational analysis. High-coverage analysis of the phosphoproteome has largely been achieved by combining pre-fractionation methods with multiple phosphopeptide enrichment approaches, at some cost in LC–MS/MS measurement time and increased sample loss. Key points for the future will be to further increase the selectivity and the recovery of enrichment methods to achieve higher sensitivity and efficiency in LC–MS/MS analysis in order to detect protein phosphorylation comprehensively, including low-abundance proteins. This is expected to lead to a more detailed understanding of the mechanisms and interactions of phosphorylation-mediated regulatory pathways in biological systems. |
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