Jump to main content
Jump to site search

Issue 23, 2019
Previous Article Next Article

An SDS-PAGE based proteomic approach for N-terminome profiling

Author affiliations


Initial sample quantity, solubilization, separation, and visualization of proteins or their proteolytically altered products are some of the challenges of the currently available solution-based N-termini enrichment methods. We therefore took advantage of the conventional SDS-PAGE system and attempted to address these challenges by proposing a simple yet reproducible, negative selection N-termini enrichment strategy coupled with mass spectrometry based protein identification. It includes in-gel protein level labeling of primary amines using d6-acetic anhydride and post-digestion negative selection of labeled N-terminal peptide(s) using N-hydroxysuccinimide activated agarose beads. We demonstrated the superiority of our method by successfully enriching protein N-termini from as low as 10 ng of bovine serum albumin. The method was validated for its applicability to a complex mixture of proteins by selectively enriching neo-N-termini generated by a site specific protease Glu-C. Its effectiveness for deep N-terminome profiling was also shown using human cell lysate. In addition, a system-wide label-free quantitative proteomic analysis of N-termini in MMP2-perturbed HCT8 cell secretome revealed substrates of several extra- and intra-cellular proteases, which are part of cell growth and proliferation and degradation pathways. In brief, the proposed method demonstrates an effective strategy not only to detect N-termini from a single protein but also for the deep and quantitative analysis of N-terminome from a limited sample amount.

Graphical abstract: An SDS-PAGE based proteomic approach for N-terminome profiling

Back to tab navigation

Supplementary files

Publication details

The article was received on 22 Aug 2019, accepted on 24 Sep 2019 and first published on 01 Oct 2019

Article type: Paper
DOI: 10.1039/C9AN01616C
Analyst, 2019,144, 7001-7009

  •   Request permissions

    An SDS-PAGE based proteomic approach for N-terminome profiling

    P. Kaushal, Y. Kwon, S. Ju and C. Lee, Analyst, 2019, 144, 7001
    DOI: 10.1039/C9AN01616C

Search articles by author