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Genetic Encoding of a Bicyclo[6.1.0]nonyne-Charged Amino Acid Enables Fast Cellular Protein Imaging by Metal-Free Ligation

Authors

  • Annika Borrmann,

    1. Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen (The Netherlands)
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    • These authors contributed equally to this work.

  • Sigrid Milles,

    1. Structural and Computational Biology Unit and Cell Biology and Biophysics Unit, EMBL, Meyerhofstrasse 1, 69117 Heidelberg (Germany)
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    • These authors contributed equally to this work.

  • Tilman Plass,

    1. Structural and Computational Biology Unit and Cell Biology and Biophysics Unit, EMBL, Meyerhofstrasse 1, 69117 Heidelberg (Germany)
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  • Jan Dommerholt,

    1. Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen (The Netherlands)
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  • Jorge M. M. Verkade,

    1. SynAffix B.V. Heyendaalsweg 135, 6525 AJ Nijmegen (The Netherlands)
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  • Prof. Dr. Manfred Wießler,

    1. Biological Chemistry, DKFZ, Im Neuenheimer Feld 280, 69120 Heidelberg (Germany)
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  • Dr. Carsten Schultz,

    1. Structural and Computational Biology Unit and Cell Biology and Biophysics Unit, EMBL, Meyerhofstrasse 1, 69117 Heidelberg (Germany)
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  • Prof. Dr. Jan C. M. van Hest,

    1. Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen (The Netherlands)
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  • Dr. Floris L. van Delft,

    Corresponding author
    1. Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen (The Netherlands)
    • Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ Nijmegen (The Netherlands)
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  • Dr. Edward A. Lemke

    Corresponding author
    1. Structural and Computational Biology Unit and Cell Biology and Biophysics Unit, EMBL, Meyerhofstrasse 1, 69117 Heidelberg (Germany)
    • Structural and Computational Biology Unit and Cell Biology and Biophysics Unit, EMBL, Meyerhofstrasse 1, 69117 Heidelberg (Germany)
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Abstract

Visualizing biomolecules by fluorescent tagging is a powerful method for studying their behaviour and function inside cells. We prepared and genetically encoded an unnatural amino acid (UAA) that features a bicyclononyne moiety. This UAA offered exceptional reactivity in strain-promoted azide–alkyne cycloadditions. Kinetic measurements revealed that the UAA reacted also remarkably fast in the inverse-electron-demand Diels–Alder cycloaddition with tetrazine-conjugated dyes. Genetic encoding of the new UAA inside mammalian cells and its subsequent selective labeling at low dye concentrations demonstrate the usefulness of the new amino acid for future imaging studies.

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