TY - CHAP
T1 - Sensitive and High-Throughput Exploration of Protein N-Termini by TMT-TAILS N-Terminomics
AU - Kalogeropoulos, Konstantinos
AU - Bundgaard, Louise
AU - auf dem Keller, Ulrich
PY - 2023
Y1 - 2023
N2 - Terminal amine isotopic labeling of substrates (TAILS) is a sensitive and robust quantitative mass spectrometry (MS)-based proteomics method used for the characterization of physiological or proteolytically processed protein N-termini, as well as other N-terminal posttranslational modifications (PTMs). TAILS is a well-established, high-throughput, negative enrichment workflow that enables system-wide exploration of N-terminomes independent of sample complexity. TAILS makes use of amine reactivity of free N-termini and a highly efficient aldehyde-functionalized polymer to deplete internal peptides generated after proteolytic digestion during sample preparation. Thereby, it enriches for natural N-termini, allowing for unbiased and complete investigation of differential proteolysis, protease substrate discovery, and analysis of N-terminal PTMs. In this chapter, we provide a state-of-the-art protocol, with detailed steps in all parts of the TAILS sample preparation, MS analysis, and post-processing of acquired data.
AB - Terminal amine isotopic labeling of substrates (TAILS) is a sensitive and robust quantitative mass spectrometry (MS)-based proteomics method used for the characterization of physiological or proteolytically processed protein N-termini, as well as other N-terminal posttranslational modifications (PTMs). TAILS is a well-established, high-throughput, negative enrichment workflow that enables system-wide exploration of N-terminomes independent of sample complexity. TAILS makes use of amine reactivity of free N-termini and a highly efficient aldehyde-functionalized polymer to deplete internal peptides generated after proteolytic digestion during sample preparation. Thereby, it enriches for natural N-termini, allowing for unbiased and complete investigation of differential proteolysis, protease substrate discovery, and analysis of N-terminal PTMs. In this chapter, we provide a state-of-the-art protocol, with detailed steps in all parts of the TAILS sample preparation, MS analysis, and post-processing of acquired data.
KW - Mass-spectrometry-based proteomics
KW - N-terminomics
KW - Degradomics
KW - Protease characterization
KW - Protease substrate discovery
KW - N-terminal posttranslational modifications
KW - TAILS
U2 - 10.1007/978-1-0716-3457-8_7
DO - 10.1007/978-1-0716-3457-8_7
M3 - Book chapter
C2 - 37665457
SN - 978-1-0716-3456-1
SN - 978-1-0716-3459-2
VL - 2718
T3 - Methods in Molecular Biology
SP - 111
EP - 135
BT - Mass Spectrometry-Based Proteomics
A2 - Gevaert, Kris
PB - Springer
ER -