Premium
Enrichment of protein N‐termini by charge reversal of internal peptides
Author(s) -
Lai Zon W.,
GomezAuli Alejandro,
Keller Eva J.,
Mayer Bettina,
Biniossek Martin L.,
Schilling Oliver
Publication year - 2015
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201500023
Subject(s) - cleavage (geology) , chemistry , posttranslational modification , acetylation , proteolysis , biochemistry , peptide , residue (chemistry) , proteomics , computational biology , biology , enzyme , paleontology , fracture (geology) , gene
Protein N‐termini provide useful information for the understanding of posttranslational processing of proteins. The majority of proteins undergo N‐terminal processing, such as proteolytic truncation or modifications like acetylation. Multiple methods currently exist for the enrichment of N‐terminal peptides for proteomic analyses. Here, we report a novel, simple, and straightforward N‐terminomic strategy, based on charge reversal of internal peptides followed by their removal through strong cation exchange chromatography. Our initial proof‐of‐concept study shows the feasibility of this technique, yielding over 3000 identifications of protein N‐termini. We further show the application of this strategy in investigating the N‐terminome of mouse embryonic fibroblasts cells deficient for both cathepsin B and L in comparison to wild type) control cells. Finally, we demonstrate that this workflow can be used in combination with a gel‐based strategy, allowing preseparation of proteins and thus providing an estimate of the molecular weight of the identified cleavage products.