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Neutron-encoded mass signatures for multiplexed proteome quantification

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Abstract

We describe a protein quantification method called neutron encoding that exploits the subtle mass differences caused by nuclear binding energy variation in stable isotopes. These mass differences are synthetically encoded into amino acids and incorporated into yeast and mouse proteins via metabolic labeling. Mass spectrometry analysis with high mass resolution (>200,000) reveals the isotopologue-embedded peptide signals, permitting quantification. Neutron encoding will enable highly multiplexed proteome analysis with excellent dynamic range and accuracy.

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Figure 1: NeuCode feasibility and scan sequence.
Figure 2: NeuCode SILAC quantitative results.

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Acknowledgements

We thank A.J. Bureta for help with figure illustrations, R. Manis for critical proofreading and A. Gasch (University of Wisconsin-Madison) for assistance in the culturing of yeast cells. This work was supported by US National Institutes of Health (NIH) grant R01 GM080148 to J.J.C. A.E.M. gratefully acknowledges support from an NIH-funded Genomic Sciences Training Program (5T32HG002760). This work was also supported by a Searle Scholars Award and Shaw Scientist award to D.J.P. and a National Science Foundation graduate fellowship and NIH training grant 5T32GM007215-37 to A.J.S.

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Author notes

  1. Alexander S Hebert and Anna E Merrill: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biomolecular Chemistry, University of Wisconsin–Madison, Madison, Wisconsin, USA

    Alexander S Hebert & Joshua J Coon

  2. Genome Center of Wisconsin, University of Wisconsin–Madison, Madison, Wisconsin, USA

    Alexander S Hebert, Anna E Merrill, Derek J Bailey, Michael S Westphall & Joshua J Coon

  3. Department of Chemistry, University of Wisconsin–Madison, Madison, Wisconsin, USA

    Anna E Merrill, Derek J Bailey, Eric R Strieter & Joshua J Coon

  4. Department of Biochemistry, University of Wisconsin–Madison, Madison, Wisconsin, USA

    Amelia J Still & David J Pagliarini

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  1. Alexander S Hebert
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  2. Anna E Merrill
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  3. Derek J Bailey
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  4. Amelia J Still
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  5. Michael S Westphall
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  6. Eric R Strieter
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  7. David J Pagliarini
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  8. Joshua J Coon
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Contributions

A.S.H. and A.E.M. designed and performed research, analyzed data and wrote the paper; D.J.B. performed theoretical calculations; M.S.W. analyzed data; E.R.S. designed research studies; D.J.P. and A.J.S. designed and prepared the murine model system; J.J.C. designed research studies and wrote the paper.

Corresponding author

Correspondence to Joshua J Coon.

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Competing interests

A.S.H. and J.J.C. are co-inventors on a patent application (US 13/660677) related in part to the material presented here.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–7 (PDF 2012 kb)

Supplementary Table 1

Myogenic differentiation protein quantitation (XLSX 219 kb)

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Hebert, A., Merrill, A., Bailey, D. et al. Neutron-encoded mass signatures for multiplexed proteome quantification. Nat Methods 10, 332–334 (2013). https://doi.org/10.1038/nmeth.2378

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