Open AccessSuper Heavy TMTpro Labeling Reagent: An Alternative and Higher-Charge-State-Amenable Stable-Isotope-Labeled TMTpro Variant
Author(s) -
Alban Ordureau,
Qing Yu,
Ryan Bomgarden,
John C. Rogers,
J. Wade Harper,
Steven P. Gygi,
João A. Paulo
Publication year - 2021
Publication title -
journal of proteome research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.644
H-Index - 161
eISSN - 1535-3907
pISSN - 1535-3893
DOI - 10.1021/acs.jproteome.0c01056
Subject(s) - reagent , peptide , chemistry , isobaric labeling , isotope , isobaric process , chromatography , label free quantification , tandem mass tag , mass spectrometry , combinatorial chemistry , quantitative proteomics , tandem mass spectrometry , biochemistry , proteomics , protein mass spectrometry , physics , quantum mechanics , gene , thermodynamics
Stable isotope labeling is a leading strategy for mass-spectrometry-based peptide quantification. Whereas TMTpro isobaric tagging can quantify up to 16 multiplexed samples in a single experiment, nonisobaric, yet chromatographically indistinguishable, variants of TMTpro reagents can be used in conjunction with the isobaric tag series for various peptide-targeting applications. Here we test the performance of two nonisobaric TMTpro variants, a stable-isotope-free TMTproZero tag and a nearly fully isotope-labeled "super-heavy" variant, shTMTpro, in a targeted assay for peptides of charge state 4+. We label each peptide with TMTproZero or Super Heavy TMTpro reagents and separately spike each peptide into a TMTpro16-labeled background (equal amount of peptide across all 16 channels). We observe that the expected 1:1 reporter ion ratio is distorted when a TMTproZero-labeled peptide is used; however, we note no such interference when shTMTpro substitutes the TMTproZero tag. Our data suggest that using the Super Heavy TMTpro reagent is an improvement over the TMTproZero reagent for the accurate quantification of high-charge-state peptides for trigger-based multiplexed assays.