Limits for Resolving Isobaric Tandem Mass Tag Reporter Ions Using Phase-Constrained Spectrum Deconvolution

Journal of Proteome Research
Christian D KelstrupJesper V Olsen

Abstract

A popular method for peptide quantification relies on isobaric labeling such as tandem mass tags (TMT), which enables multiplexed proteome analyses. Quantification is achieved by reporter ions generated by fragmentation in a tandem mass spectrometer. However, with higher degrees of multiplexing, the smaller mass differences between the reporter ions increase the mass resolving power requirements. This contrasts with faster peptide sequencing capabilities enabled by lowered mass resolution on Orbitrap instruments. It is therefore important to determine the mass resolution limits for highly multiplexed quantification when maximizing proteome depth. Here, we defined the lower boundaries for resolving TMT reporter ions with 0.0063 Da mass differences using an ultra-high-field Orbitrap mass spectrometer. We found the optimal method depends on the relative ratio between closely spaced reporter ions and that 64 ms transient acquisition time provided sufficient resolving power for separating TMT reporter ions with absolute ratio changes up to 16-fold. Furthermore, a 32 ms transient processed with phase-constrained spectrum deconvolution provides >50% more identifications with >99% quantified but with a slight loss in quantification pre...Continue Reading

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Oct 29, 2014·Journal of Proteome Research·Christian D KelstrupJesper V Olsen
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Dec 1, 2017·Journal of Proteome Research·Christian D KelstrupJesper V Olsen
Jan 10, 2018·Angewandte Chemie·Michael StadlmeierThomas Carell

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Citations

May 23, 2020·Nature Metabolism·Khanh-Van TranSøren Nielsen
Feb 14, 2020·Molecular & Cellular Proteomics : MCP·Dorte B Bekker-JensenJesper V Olsen
Jan 22, 2020·Nature Biotechnology·Jessica PerrinGiovanna Bergamini
Jan 17, 2021·Molecular & Cellular Proteomics : MCP·Dorte B Bekker-JensenJesper V Olsen
Apr 7, 2020·Analytical Chemistry·Qing YuDevin K Schweppe
Aug 21, 2021·Chemical Reviews·Sem TamaraAlbert J R Heck
Dec 31, 2021·Analytical Chemistry·Claudia CtorteckaKarl Mechtler

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