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A novel thermostable protein-tag: optimization of the Sulfolobus solfataricus DNA- alkyl-transferase by protein engineering

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An Erratum to this article was published on 14 December 2015

Abstract

In the last decade, a powerful biotechnological tool for the in vivo and in vitro specific labeling of proteins (SNAP-tag™ technology) was proposed as a valid alternative to classical protein-tags (green fluorescent proteins, GFPs). This was made possible by the discovery of the irreversible reaction of the human alkylguanine-DNA-alkyl-transferase (hAGT) in the presence of benzyl-guanine derivatives. However, the mild reaction conditions and the general instability of the mesophilic SNAP-tag™ make this new approach not fully applicable to (hyper-)thermophilic and, in general, extremophilic organisms. Here, we introduce an engineered variant of the thermostable alkylguanine-DNA-alkyl-transferase from the Archaea Sulfolobus solfataricus (SsOGT-H5), which displays a catalytic efficiency comparable to the SNAP-tag™ protein, but showing high intrinsic stability typical of proteins from this organism. The successful heterologous expression obtained in a thermophilic model organism makes SsOGT-H5 a valid candidate as protein-tag for organisms living in extreme environments.

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Acknowledgments

We gratefully thank Castrese Morrone for help in some experiments. This work was supported by: (1) Short Term Mobility Program of the National Research Council of Italy; (2) FIRB-Futuro in Ricerca RBFR12OO1G_002-Nematic and Merit RBNE08YFN3-Molecular Oncology; (3) BIO2013-44953-R Project.

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    Authors and Affiliations

    1. Institute of Biosciences and BioResources, National Council of Research of Italy, Via P. Castellino 111, 80131, Naples, Italy

      Antonella Vettone, Mario Serpe, Giovanni del Monaco, Anna Valenti, Mosé Rossi, Maria Ciaramella & Giuseppe Perugino

    2. Centro De Biología Molecular Severo Ochoa, Nicolás Cabrera 1, Cantoblanco, 28049, Madrid, Spain

      Aurelio Hidalgo & José Berenguer

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    1. Antonella Vettone
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    2. Mario Serpe
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    3. Aurelio Hidalgo
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    4. José Berenguer
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    5. Giovanni del Monaco
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    6. Anna Valenti
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    7. Mosé Rossi
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    8. Maria Ciaramella
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    9. Giuseppe Perugino
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    Correspondence to Giuseppe Perugino.

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    Communicated by S. Albers.

    A. Vettone and M. Serpe equally contributed to the present work.

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    Vettone, A., Serpe, M., Hidalgo, A. et al. A novel thermostable protein-tag: optimization of the Sulfolobus solfataricus DNA- alkyl-transferase by protein engineering. Extremophiles 20, 1–13 (2016). https://doi.org/10.1007/s00792-015-0791-9

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