Real-time monitoring of cell-free translation on a quartz-crystal microbalance

Shuntaro Takahashi, Masaaki Iida, Hiroyuki Furusawa, Yoshihiro Shimizu, Takuya Ueda, Yoshio Okahata

研究成果: Article

18 引用 (Scopus)

抄録

The efficiency of protein synthesis is often regulated post- transcriptionally by sequences within the mRNA. To investigate the reactions of protein translation, we established a system that allowed real-time monitoring of protein synthesis using a cell-free translation mixture and a 27 MHz quartz-crystal microbalance (QCM). Using an mRNA that encoded a fusion polypeptide comprising the streptavidin-binding peptide (SBP) tag, a portion of Protein D as a spacer, and the SecM arrest sequence, we could follow the binding of the SBP tag, while it was displayed on the 70S ribosome, to a streptavidin-modified QCM over time. Thus, we could follow a single turnover of protein synthesis as a change in mass. This approach allowed us to evaluate the effects of different antibiotics and mRNA sequences on the different steps of translation. From the results of this study, we have determined that both the formation of the initiation complex from the 70S ribosome, mRNA, and fMet-tRNAfMet and the accommodation of the second aminoacyl-tRNA to the initiation complex are rate-limiting steps in protein synthesis.

元の言語English
ページ(範囲)9326-9332
ページ数7
ジャーナルJournal of the American Chemical Society
131
発行部数26
DOI
出版物ステータスPublished - 2009 7 8
外部発表Yes

Fingerprint

Quartz Crystal Microbalance Techniques
Quartz crystal microbalances
Proteins
Monitoring
Messenger RNA
Ribosomes
Peptides
Streptavidin
Computer Systems
Protein Biosynthesis
Transfer RNA
Polypeptides
Antibiotics
Anti-Bacterial Agents
Fusion reactions

ASJC Scopus subject areas

  • Chemistry(all)
  • Catalysis
  • Biochemistry
  • Colloid and Surface Chemistry

これを引用

Takahashi, S., Iida, M., Furusawa, H., Shimizu, Y., Ueda, T., & Okahata, Y. (2009). Real-time monitoring of cell-free translation on a quartz-crystal microbalance. Journal of the American Chemical Society, 131(26), 9326-9332. https://doi.org/10.1021/ja9019947

Real-time monitoring of cell-free translation on a quartz-crystal microbalance. / Takahashi, Shuntaro; Iida, Masaaki; Furusawa, Hiroyuki; Shimizu, Yoshihiro; Ueda, Takuya; Okahata, Yoshio.

:: Journal of the American Chemical Society, 巻 131, 番号 26, 08.07.2009, p. 9326-9332.

研究成果: Article

Takahashi, S, Iida, M, Furusawa, H, Shimizu, Y, Ueda, T & Okahata, Y 2009, 'Real-time monitoring of cell-free translation on a quartz-crystal microbalance', Journal of the American Chemical Society, 巻. 131, 番号 26, pp. 9326-9332. https://doi.org/10.1021/ja9019947
Takahashi, Shuntaro ; Iida, Masaaki ; Furusawa, Hiroyuki ; Shimizu, Yoshihiro ; Ueda, Takuya ; Okahata, Yoshio. / Real-time monitoring of cell-free translation on a quartz-crystal microbalance. :: Journal of the American Chemical Society. 2009 ; 巻 131, 番号 26. pp. 9326-9332.
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