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Widespread bacterial protein histidine phosphorylation revealed by mass spectrometry-based proteomics

Nature Methods volume 15, pages 187190 (2018) | Download Citation

Abstract

For decades, major difficulties in analyzing histidine phosphorylation have limited the study of phosphohistidine signaling. Here we report a method revealing widespread and abundant protein histidine phosphorylation in Escherichia coli. We generated an extensive E. coli phosphoproteome data set, in which a remarkably high percentage (10%) of phosphorylation sites are phosphohistidine sites. This resource should help enable a better understanding of the biological function of histidine phosphorylation.

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Acknowledgements

We thank R. Zheng and B. Kuster of the Technical University Munich, Freising, Germany, for providing us with synthetic peptides used in this study. This work was supported by the Roadmap Initiative Proteins@Work funded by the Netherlands Organization for Scientific Research (NWO) (to A.J.R.H.; project number 184.032.201) and by the MSMed program, funded by the European Union's Horizon 2020 Framework Programme (to A.J.R.H.; grant agreement number 686547). S.L. acknowledges support from the Netherlands Organization for Scientific Research (NWO) through a VIDI grant (project 723.013.008).

Author information

Author notes

    • Clement M Potel
    •  & Miao-Hsia Lin

    These authors contributed equally to this work.

Affiliations

  1. Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Utrecht, the Netherlands.

    • Clement M Potel
    • , Miao-Hsia Lin
    • , Albert J R Heck
    •  & Simone Lemeer
  2. Netherlands Proteomics Center, Utrecht, the Netherlands.

    • Clement M Potel
    • , Miao-Hsia Lin
    • , Albert J R Heck
    •  & Simone Lemeer

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Contributions

C.M.P., M.-H.L., A.J.R.H. and S.L. designed the experiment. C.M.P. and M.-H.L. performed the experiments. C.M.P., M.-H.L. and S.L. performed data analysis. C.M.P., M.-H.L., A.J.R.H. and S.L. wrote the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Albert J R Heck or Simone Lemeer.

Integrated supplementary information

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–12 and Supplementary Tables 4 and 6.

  2. 2.

    Life Sciences Reporting Summary

Excel files

  1. 1.

    Supplementary Table 1

    List of identified phosphohistidine sites from chemically phosphorylated protein mixtures.

  2. 2.

    Supplementary Table 2

    List of all identified phosphosites (STYH) from E. coli, including score, localization probability and precursor intensity in the different growth conditions.

  3. 3.

    Supplementary Table 3

    Comparison of identified phosphohistidine sites from antibody- and MS-based approaches.

  4. 4.

    Supplementary Table 5

    List of quantified phosphosites (STYH) and statistical analysis of detected changes between exponential and stationary phases of glycerol-fed E. coli.

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DOI

https://doi.org/10.1038/nmeth.4580

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