Journal article
Neutron-Encoded Mass Signatures for Quantitative Top-Down Proteomics
Analytical chemistry (Washington), Vol.86(5), pp.2314-2319
03/04/2014
DOI: 10.1021/ac403579s
PMCID: PMC3983007
PMID: 24475910
Abstract
The ability to acquire highly accurate
quantitative data is an
increasingly important part of any proteomics experiment, whether
shotgun or top-down approaches are used. We recently developed a quantitation
strategy for peptides based on neutron encoding, or NeuCode SILAC,
which uses closely spaced heavy isotope-labeled amino acids and high-resolution
mass spectrometry to provide quantitative data. We reasoned that the
strategy would also be applicable to intact proteins and could enable
robust, multiplexed quantitation for top-down experiments. We used
yeast lysate labeled with either
13
C
6
15
N
2
-lysine or
2
H
8
-lysine, isotopologues
of lysine that are spaced 36 mDa apart. Proteins having such close
spacing cannot be distinguished during a medium resolution scan, but
upon acquiring a high-resolution scan, the two forms of the protein
with each amino acid are resolved and the quantitative information
revealed. An additional benefit NeuCode SILAC provides for top down
is that the spacing of the isotope peaks indicates the number of lysines
present in the protein, information that aids in identification. We
used NeuCode SILAC to quantify several hundred isotope distributions,
manually identify and quantify proteins from 1:1, 3:1, and 5:1 mixed
ratios, and demonstrate MS
2
-based quantitation using ETD.
Details
- Title: Subtitle
- Neutron-Encoded Mass Signatures for Quantitative Top-Down Proteomics
- Creators
- Timothy W Rhoads - Department of ChemistryChristopher M Rose - Department of ChemistryDerek J Bailey - Department of ChemistryNicholas M Riley - Department of ChemistryRosalynn C Molden - Department of Biochemistry and BiophysicsAmelia J Nestler - Department of ChemistryAnna E Merrill - Department of ChemistryLloyd M Smith - Department of ChemistryAlexander S Hebert - Department of ChemistryMichael S Westphall - Department of ChemistryDavid J Pagliarini - Department of ChemistryBenjamin A Garcia - Department of Biochemistry and BiophysicsJoshua J Coon - Department of Chemistry
- Resource Type
- Journal article
- Publication Details
- Analytical chemistry (Washington), Vol.86(5), pp.2314-2319
- DOI
- 10.1021/ac403579s
- PMID
- 24475910
- PMCID
- PMC3983007
- NLM abbreviation
- Anal Chem
- ISSN
- 0003-2700
- eISSN
- 1520-6882
- Publisher
- American Chemical Society
- Language
- English
- Date published
- 03/04/2014
- Academic Unit
- Pathology; Injury Prevention Research Center
- Record Identifier
- 9984047975702771
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