Multiplexed protein quantitation in Saccharomyces cerevisiae using amine-reactive isobaric tagging reagents
Open accessAbstract
We describe here a multiplexed protein quantitation strategy that provides relative and absolute measurements of proteins in complex mixtures. At the core of this methodology is a multiplexed set of isobaric reagents that yield amine-derivatized peptides. The derivatized peptides are indistinguishable in MS, but exhibit intense low-mass MS/MS signature ions that support quantitation. In this study, we have examined the global protein expression of a wild-type yeast strain and the isogenic upf1Delta and xrn1Delta mutant strains that are defective in the nonsense-mediated mRNA decay and the general 5' to 3' decay pathways, respectively. We also demonstrate the use of 4-fold multiplexing to enable relative protein measurements simultaneously with determination of absolute levels of a target protein using synthetic isobaric peptide standards. We find that inactivation of Upf1p and Xrn1p causes common as well as unique effects on protein expression.
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Chemical labelling has become a widely employed approach not only in NPs research, but also in proteomics, food-safety, forensics, and environmental analysis.
“Chemical labelling has become a widely employed approach not only in the NPs research, but also in other research fields including proteomics,98,99 food-safety,100,101 forensics,102,103 environmental analysis.104,105”
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Chemical labeling is in principle universally applicable, but because labels are introduced later in sample processing, some robustness advantages are lost.
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From Accurate proteome-wide label-free quantification by delayed normalization and maximal peptide ratio extraction, termed MaxLFQ (2014) · cites this paper as [40]
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Self-citation
- Authors
- Ross, P. L., Huang, Y. N., Marchese, J. N., Williamson, B., Parker, K., Hattan, S., Khainovski, N., Pillai, S., Dey, S., Daniels, S., Purkayastha, S., Juhasz, P., Martin, S., Bartlet-Jones, M., He, F., Jacobson, A., & Pappin, D. J.
- Affiliations
- Biocompatibles (United Kingdom) (GB)
- University of Massachusetts Chan Medical School (US)
- Imperial College London (GB)
- Corresponding author
- Pappin, Darryl J.
- Venue
- Molecular & Cellular Proteomics
- Publisher
- Elsevier BV
- Published
- 2004-12
- Volume
- 3
- Issue
- 12
- Pages
- 1154-1169
- ISSN
- 1535-9476 (ISSN-L)
- OpenAlex
- W2120016245
- Cited by
- 4,401
- References
- 36
- Citations by year
- 2026: 47 · 2025: 61 · 2024: 107 · 2023: 99 · 2022: 108 · 2021: 129 · 2020: 139 · 2019: 167 · 2018: 198 · 2017: 219 · 2016: 249 · 2015: 260 · 2014: 312 · 2013: 335 · 2012: 374
- FWCI
- 65.55
- Type
- journal article
- Language
- en
- Open access
- hybrid
- doi.org
- Licence
- Locations
- Molecular & Cellular Proteomics (publishedVersion, cc-by)
- PubMed (publishedVersion)
- Cold Spring Harbor Laboratory Institutional Repository (Cold Spring Harbor Laboratory) (acceptedVersion)
- Funders
- National Institutes of Health
- National Institute of General Medical Sciences (GM27757, R37GM027757, R01GM027757)
- Topics
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ubiquitin and proteasome pathways
- Keywords
- peptide derivatization, nonsense-mediated mRNA decay, Saccharomyces cerevisiae, proteomics
- MeSH terms
- Cations, Chromatography, Ion Exchange, Chromatography, Liquid, Exoribonucleases, Fungal Proteins, Indicators and Reagents, Ions, Models, Chemical, Peptides, Phenotype, RNA, Messenger, Saccharomyces cerevisiae, Mass Spectrometry, Succinimides, Down-Regulation, RNA Helicases, Saccharomyces cerevisiae Proteins, Proteomics
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