Recently, Analytical Chemistry, an international academic journal, published "Multi-in-One: Multiple-Proteases, One-Hour-Shot Strategy for Fast and High-Coverage Phosphoproteomic Investigation", the research results of Zeng Rong's research group from Key Laboratory of Systems Biology, HIAS/Centerfor Excellence in Molecular Cell Science, Chinese Academy of Sciences. This study combined multiple proteolytic enzymes and data apathy collection patterns for the first time, and proposed a new proteome research method-"Multi-in-One".
Overview of the "Multi-in-One" Strategy.
Based on mass spectrometry, large-scale quantitative proteomics has broad application prospects in biomedical research, but the semi-randomness of traditional data-dependent acquisition (DDA) and the preference of single trypsin hydrolases have limitations in large-scale analysis of low-abundance proteins and modification sites.
In order to overcome this problem, Zeng Rong's research group made full use of the complementarity of different protease digestions and high repeatability of data-independent acquisition (DIA), which greatly improved the coverage of protein and phosphorylation modification sequences and enhanced the identification and quantitative repeatability between samples. The one-step enrichment and one-stop data collection of the method simplify the experimental process to the greatest extent and enhance the operability and practicability of the method. Compared with the traditional method, the new method has improved the ability to identify phosphosites by 60%, shortened the data acquisition time by 95%, and improved the experimental repeatability by more than 100%. Researchers applied the Multi-in-One method to the study of early activated brown adipocytes and found that this method has unique advantages in quantifying phosphorylation modification sites of low-abundance transcription factors. Highly complementary hydrolases can discover and quantify some phosphorylation modification sites that are not easily covered by trypsin, thus revealing some new regulated kinase substrate sequences. This approach provides a novel strategy for rapid high-coverage proteome quantification that can be widely applied to deep and large-scale proteome and its modification analysis.
Under the joint guidance of research fellow Zeng Rong and associate research fellow Li Qingrun, this work was completed by Gao Xiaojing, a doctoral student from Zeng Rong's research group, and was strongly supported by Professor Liu Yansheng from Yale Cancer Biology Institute, Yale School of Medicine. This work was funded by the Strategic Priority Research Program of the Chinese Academy of Sciences, the National Key Research and Development Program of the Ministry of Science and Technology of the People's Republic of China, and the Natural Science Foundation of China.
Article Link:https://pubs.acs.org/doi/10.1021/acs.analchem.0c00906
Editor | Jiang Xuchen
