4.7 Article

PBC, an easy and efficient strategy for high-throughput protein C-terminome profiling

期刊

出版社

FRONTIERS MEDIA SA
DOI: 10.3389/fcell.2022.995590

关键词

C-terminomics; chemical derivatization; enrichment; high-efficiency; post-translation modification (PTM)

资金

  1. National Key R and D Program of China [2020YFE0202200]
  2. Natural Science Foundation of China [32071432, 21877062, 32171434]
  3. Basic research projects of Shanghai Science and Technology Commission science and technology innovation action plan [19JC1416300]
  4. open fund of state key laboratory of Pharmaceutical Biotechnology, Nanjing University, China [KF-202201]
  5. NSFC-NRF (China-Korea) Joint Research Program [82111540276]

向作者/读者索取更多资源

High-throughput profiling of protein C-termini is challenging, but this study developed a strategy using 2-PCA and biotin labeling for efficient and high-throughput analysis of protein C-terminome.
High-throughput profiling of protein C-termini is still a challenging task. Proteomics provides a powerful technology for systematic and high-throughput study of protein C-termini. Various C-terminal peptide enrichment strategies based on chemical derivatization and chromatography separation have been reported. However, they are still costly and time-consuming, with low enrichment efficiency for C-terminal peptides. In this study, by taking advantage of the high reaction selectivity of 2-pyridinecarboxaldehyde (2-PCA) with an alpha-amino group on peptide N-terminus and high affinity between biotin and streptavidin, we developed a 2-PCA- and biotin labeling-based C-terminomic (PBC) strategy for a high-efficiency and high-throughput analysis of protein C-terminome. Triplicates of PBC experiments identified a total of 1,975 C-terminal peptides corresponding to 1,190 proteins from 293 T cell line, which is 180% higher than the highest reported number of C-terminal peptides identified from mammalian cells by chemical derivatization-based C-terminomics study. The enrichment efficiency (68%) is the highest among the C-terminomics methods currently reported. In addition, we not only uncovered 50 proteins with truncated C-termini which were significantly enriched in extracellular exosome, vesicle, and ribosome by a bioinformatic analysis but also systematically characterized the whole PTMs on C-terminal in 293 T cells, suggesting PBC as a powerful tool for protein C-terminal degradomics and PTMs investigation. In conclusion, the PBC strategy would benefit high-efficiency and high-throughput profiling of protein C-terminome.

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