A protocol for analyzing the protein terminome of human cancer cell line culture supernatants

Kazuya Tsumagari; Chih-Hsiang Chang; Yasushi Ishihama

doi:10.1016/j.xpro.2021.100682

A protocol for analyzing the protein terminome of human cancer cell line culture supernatants

STAR Protoc. 2021 Jul 22;2(3):100682. doi: 10.1016/j.xpro.2021.100682. eCollection 2021 Sep 17.

Authors

Kazuya Tsumagari^{1

2}, Chih-Hsiang Chang¹, Yasushi Ishihama^{1

3}

Affiliations

¹ Department of Molecular and Cellular BioAnalysis, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto 606-8501, Japan.
² Center for Integrated Medical Research, Keio University School of Medicine, Tokyo 160-8582, Japan.
³ Laboratory of Clinical and Analytical Chemistry, National Institute of Biomedical Innovation, Health and Nutrition, Ibaraki, Osaka 567-0085, Japan.

Abstract

Characterization of protein termini is essential for understanding how the proteome is generated through biological processes such as post-translational proteolytic events. Here, we introduce a practical protocol for terminomics to achieve simple and sensitive N- and C-terminal peptide enrichment. We apply it to the terminome analysis of culture supernatants of a human cancer cell line for the purpose of identifying ectodomain shedding substrate cleavage sites with 10 μg protein per sample. For complete details on the use and execution of this protocol, please refer to Tsumagari et al. (2021).

Keywords: Cell Membrane; Mass Spectrometry; Protein Biochemistry; Proteomics; Systems biology.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cell Line, Tumor / metabolism
Humans
Isotope Labeling / methods*
Peptide Hydrolases / metabolism
Peptides / chemistry
Protein Domains / physiology*
Protein Processing, Post-Translational / physiology
Proteolysis / drug effects
Proteome / analysis
Proteomics / methods*
Tandem Mass Spectrometry / methods

Substances

Peptides
Proteome
Peptide Hydrolases