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Cell-free translation reconstituted with purified components

Nature Biotechnology volume 19, pages 751–755 (2001)Cite this article

Abstract

We have developed a protein-synthesizing system reconstituted from recombinant tagged protein factors purified to homogeneity. The system was able to produce protein at a rate of about 160 μg/ml/h in a batch mode without the need for any supplementary apparatus. The protein products were easily purified within 1 h using affinity chromatography to remove the tagged protein factors. Moreover, omission of a release factor allowed efficient incorporation of an unnatural amino acid using suppressor transfer RNA (tRNA).

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Acknowledgements

We thank S. Yokoyama, The University of Tokyo, for providing the expression plasmid encoding the ArgRS gene. This work was supported by a grant (JSPS-RFTF 96100306) from The Japan Society for the Promotion of Science (to T.U.).

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Authors and Affiliations

  1. Department of Chemistry and Biotechnology, Graduate School of Engineering, The University of Tokyo, Bldg FSB401, 5-1-5, Kasiwanoha, Kashiwa, 277-8562, Chiba prefecture, Japan

    Yoshihiro Shimizu & Yukihide Tomari

  2. Department of Integrated Biosciences, Graduate School of Frontier Sciences, The University of Tokyo, Bldg FSB401, 5-1-5, Kasiwanoha, Kashiwa, 277-8562, Chiba prefecture, Japan

    Akio Inoue, Tsutomu Suzuki & Takuya Ueda

  3. Department of Biomolecular Science, Faculty of Engineering, Gifu University, Yanagido, 501-1193, Gifu, Japan

    Takashi Yokogawa & Kazuya Nishikawa

Authors
  1. Yoshihiro Shimizu
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  4. Tsutomu Suzuki
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The online version of the original article can be found at 10.1038/90773

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Shimizu, Y., Inoue, A., Tomari, Y. et al. Cell-free translation reconstituted with purified components. Nat Biotechnol 19, 751–755 (2001). https://doi.org/10.1038/90802

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