A Rapid Approach for Isobaric Peptide Termini Labeling
Isobaric peptide termini labeling (IPTL) is a recently introduced approach to the chemical labeling of peptides with isotopic reagents. Peptides derived from two different samples are labeled at the N terminus and at the C terminus with isotopically labeled reagents that have identical mass differences. To obtain isobaric peptides, labeling is carried out such that the introduced mass increase at one terminus will exactly match the mass decrease at the other terminus (and the other way around). This results in product ion spectra that display the quantitative difference of the peptide signal derived from the two samples for every b-ion and y-ion in the spectrum. The original IPTL approach required the selective modification of lysines followed by C-18 micropurification of modified peptides and reaction of the N termini. Here, we describe a new approach for IPTL that is based on the selective modification of the peptide N termini with succinic anhydride and subsequent reductive amination of C-terminal lysines with formaldehyde and cyanoborohydride. Both reactions can be carried out in one pot within 10 min and without C-18 micropurification. In addition, we present the software package IsobariQ for straightforward data analysis.
Key wordsChemical labeling Dimethylation Isobaric labeling IsobariQ Isobaric peptide termini labeling iTRAQ Mass spectrometry Quantitative proteomics Succinylation TMT
IPTL was supported by the National Program for Research in Functional Genomics in Norway (FUGE, project no. 183418/S10) of the Norwegian Research Council, MLSUiO and FUGE-Øst to B.T.
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