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Detection of post-translational modifications in single peptides using electron tunnelling currents

Nature Nanotechnology volume 9, pages 835–840 (2014)Cite this article

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Abstract

Post-translational modifications alter the properties of proteins through the cleavage of peptide bonds or the addition of a modifying group to one or more amino acids1,2,3,4,5,6,7. These modifications allow proteins to perform their primary biological functions8,9,10,11, but single-protein studies of post-translational modifications have been hindered by a lack of suitable analysis methods12,13,14,15,16. Here, we show that single amino acids can be identified using electron tunnelling currents measured as the individual molecules pass through a nanoscale gap between electrodes. We identify 12 different amino acids and the post-translational modification phosphotyrosine, which is involved in the process that switches enzymes on and off17,18,19,20. Furthermore, we show that the conductance measurements can be used to partially sequence peptides of an epidermal growth factor receptor substrate, and can discriminate a peptide from its phosphorylated variant.

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Figure 1: Single-molecule proteomic analysis of post-translational modification, using tunnelling currents conducted via single amino-acid molecules.
Figure 2: Single-molecule identification of amino-acid and phosphotyrosine molecules.
Figure 3: Discrimination of individual tryosine (Y) and phosphorylated tyrosine (pY).
Figure 4: Single-molecule peptide identification and discrimination of post-translational modification.

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Acknowledgements

This research was partially supported by the Japan Society for the Promotion of Science (JSPS) through its Funding Program for World-Leading Innovative R&D on Science and Technology, and KAKENHI grant no. 26220603.

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Authors and Affiliations

  1. The Institute of Scientific and Industrial Research, Osaka University, 8-1 Mihogaoka, Ibaraki, Osaka, 567-0047, Japan

    Takahito Ohshiro, Makusu Tsutsui, Kazumichi Yokota, Masayuki Furuhashi, Masateru Taniguchi & Tomoji Kawai

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  1. Takahito Ohshiro
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  2. Makusu Tsutsui
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  3. Kazumichi Yokota
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  4. Masayuki Furuhashi
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  5. Masateru Taniguchi
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  6. Tomoji Kawai
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Contributions

M.Ta. and T.K. planned and designed the experiments. T.O., M.Ts., K.Y. and M.F. participated in fabrications of nano-MCBJs and single-molecule detection measurements. T.O., M.Ts., K.Y. and M.Ta. performed data analyses. T.O., M.Ta. and T.K. co-wrote the paper.

Corresponding authors

Correspondence to Masateru Taniguchi or Tomoji Kawai.

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The authors declare no competing financial interests.

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Ohshiro, T., Tsutsui, M., Yokota, K. et al. Detection of post-translational modifications in single peptides using electron tunnelling currents. Nature Nanotech 9, 835–840 (2014). https://doi.org/10.1038/nnano.2014.193

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