pr1c00833_si_001.pdf (298.47 kB)
Retention Time Prediction for TMT-Labeled Peptides in Proteomic LC-MS Experiments
journal contribution
posted on 2022-04-01, 20:39 authored by Benilde Mizero, Carina Villacrés, Victor Spicer, Rosa Viner, Julian Saba, Bhavinkumar Patel, Sergei Snovida, Penny Jensen, Andreas Huhmer, Oleg V. KrokhinWe present the first detailed study
of chromatographic behavior
of peptides labeled with tandem mass tags (TMT and TMTpro) in 2D LC
for proteomic applications. Carefully designed experimental procedures
have permitted generating data sets of over 100,000 nonlabeled and
TMT-labeled peptide pairs for the low pH RP in the second separation
dimension and data sets of over 10,000 peptide pairs for high-pH RP,
HILIC (amide and silica), and SCX separations in the first separation
dimension. The average increase in peptide RPLC (0.1% formic acid)
retention upon TMT labeling was found to be 3.3% acetonitrile (linear
water/acetonitrile gradients), spanning a range of −4 to 10.3%.
In addition to the bulk peptide properties such as length, hydrophobicity,
and the number of labeled residues, we found several sequence-dependent
features mostly associated with differences in N-terminal chemistry.
The behavior of TMTpro-labeled peptides was found to be very similar
except for a slightly higher hydrophobicity: an average retention
shift of 3.7% acetonitrile. The respective versions of the sequence-specific
retention calculator (SSRCalc) model have been developed to accommodate
both TMT chemistries, showing identical prediction accuracy (R2 ∼ 0.98) for labeled and nonlabeled
peptides. Higher retention for TMT-labeled peptides was observed for
high-pH RP and HILIC separations, while SCX selectivity remained virtually
unchanged.
History
Usage metrics
Categories
Keywords
tandem mass tagssecond separation dimensionfirst separation dimensionfirst detailed studyspecific retention calculatorretention time predictionbulk peptide propertiesaverage retention shift000 peptide pairslow ph rp2 suplabeled peptide pairsslightly higher hydrophobicityfound several sequencehigher retentionph rppeptide rplcr average increase>< sup000 nonlabeled− 4terminal chemistrysimilar exceptsilica ),scx separationsrespective versionsproteomic lcproteomic applicationspeptides labelednonlabeled peptidesms experimentslinear waterlabeled residueslabeled peptidesformic aciddata sets2d lc