Cell-free translation reconstituted with purified components

Yoshihiro Shimizu; Akio Inoue; Yukihide Tomari; Tsutomu Suzuki; Takashi Yokogawa; Kazuya Nishikawa; Takuya Ueda

doi:10.1038/90802

Cell-free translation reconstituted with purified components

Yoshihiro Shimizu, Akio Inoue, Yukihide Tomari, Tsutomu Suzuki, Takashi Yokogawa, Kazuya Nishikawa, Takuya Ueda^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

1062 Citations (Scopus)

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).

Original language	English
Pages (from-to)	751-755
Number of pages	5
Journal	Nature biotechnology
Volume	19
Issue number	8
DOIs	https://doi.org/10.1038/90802
Publication status	Published - 2001
Externally published	Yes

ASJC Scopus subject areas

Biotechnology
Bioengineering
Applied Microbiology and Biotechnology
Molecular Medicine
Biomedical Engineering

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10.1038/90802

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title = "Cell-free translation reconstituted with purified components",

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).",

author = "Yoshihiro Shimizu and Akio Inoue and Yukihide Tomari and Tsutomu Suzuki and Takashi Yokogawa and Kazuya Nishikawa and Takuya Ueda",

note = "Funding Information: Note: Supplementary information can be found on the Nature Biotechnology website in Web Extras (http://biotech.nature.com/web_extras) Acknowledgments 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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doi = "10.1038/90802",

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T1 - Cell-free translation reconstituted with purified components

AU - Shimizu, Yoshihiro

AU - Inoue, Akio

AU - Tomari, Yukihide

AU - Suzuki, Tsutomu

AU - Yokogawa, Takashi

AU - Nishikawa, Kazuya

AU - Ueda, Takuya

N1 - Funding Information: Note: Supplementary information can be found on the Nature Biotechnology website in Web Extras (http://biotech.nature.com/web_extras) Acknowledgments 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.).

PY - 2001

Y1 - 2001

N2 - 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).

AB - 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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Cell-free translation reconstituted with purified components

Abstract

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