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Site-Specific Incorporation of Fluorescent Nonnatural Amino Acids into Proteins and Its Application to Fluorescence Analysis of Proteins

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Book cover Molecular Science of Fluctuations Toward Biological Functions

Abstract

Nonnatural amino acid mutagenesis using expanded genetic codes allows us to introduce artificial functional groups such as fluorophores at specific sites of proteins. By applying this technique, site-specific fluorescence labeling has been achieved. This method enables to generate fluorescent protein probes that show ligand-dependent fluorescence change based on fluorescence quenching of the incorporated fluorophores by endogenous Trp residues. This strategy has been proved to be effective using ligand-binding proteins and antibodies. Moreover, site-specific double-labeling of ligand-binding proteins with two fluorophores using two expanded genetic codons has achieved ratiometric detection of the ligand binding based on FRET and fluorescence quenching. These achievements demonstrate the usefulness of the site-specific incorporation of fluorescent nonnatural amino acids and their potential utility as research and diagnostic tools.

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Acknowledgments

The author deeply appreciates the contributions of all the coworkers in this research project. This work was partly supported by a Grant-in-Aid for Scientific Research on Innovative Areas (20107005) from the Ministry of Education, Culture, Sports, Science, and Technology, Japan.

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Authors and Affiliations

  1. School of Materials Science, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa, 923-1292, Japan

    Takahiro Hohsaka

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  1. Takahiro Hohsaka
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Correspondence to Takahiro Hohsaka .

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Editors and Affiliations

  1. Kyoto University, Kyoto, Japan

    Masahide Terazima

  2. Nara Institute of Science and Tech, Ikoma, Japan

    Mikio Kataoka

  3. Sojo University, Kumamoto, Japan

    Ryuichi Ueoka

  4. Nagoya University, Nagoya, Japan

    Yuko Okamoto

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Hohsaka, T. (2016). Site-Specific Incorporation of Fluorescent Nonnatural Amino Acids into Proteins and Its Application to Fluorescence Analysis of Proteins. In: Terazima, M., Kataoka, M., Ueoka, R., Okamoto, Y. (eds) Molecular Science of Fluctuations Toward Biological Functions . Springer, Tokyo. https://doi.org/10.1007/978-4-431-55840-8_5

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