Cell-free cotranslation and selection using in vitro virus for high-throughput analysis of protein-protein interactions and complexes

Etsuko Miyamoto-Sato, Masamichi Ishizaka, Kenichi Horisawa, Seiji Tateyama, Hideaki Takashima, Shinichiro Fuse, Kaori Sue, Naoya Hirai, Kazuyo Masuoka, Hiroshi Yanagawa

研究成果: ジャーナルへの寄稿記事

37 引用 (Scopus)

抄録

We have developed a simple and totally in vitro selection procedure based on cell-free cotranslation using a highly stable and efficient in vitro virus (IVV). Cell-free cotranslation of tagged bait and prey proteins is advantageous for the formation of protein complexes and allows high-throughput analysis of protein-protein interactions (PPI) as a result of providing in vitro instead of in vivo preparation of bait proteins. The use of plural selection rounds and a two-step purification of the IVV selection, followed by in vitro post-selection, is advantageous for decreasing false positives. In a single experiment using bait Fos, more than 10 interactors, including not only direct, but also indirect interactions, were enriched. Further, previously unidentified proteins containing novel leucine zipper (L-ZIP) motifs with minimal binding sites identified by sequence alignment as functional elements were detected as a result of using a randomly primed cDNA library. Thus, we consider that this simple IVV selection system based on cell-free cotranslation could be applicable to high-throughput and comprehensive analysis of PPI and complexes in large-scale settings involving parallel bait proteins.

元の言語英語
ページ(範囲)710-717
ページ数8
ジャーナルGenome Research
15
発行部数5
DOI
出版物ステータス出版済み - 5 1 2005

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Viruses
Proteins
In Vitro Techniques
Leucine Zippers
Sequence Alignment
Gene Library
Binding Sites

All Science Journal Classification (ASJC) codes

  • Genetics
  • Genetics(clinical)

これを引用

Cell-free cotranslation and selection using in vitro virus for high-throughput analysis of protein-protein interactions and complexes. / Miyamoto-Sato, Etsuko; Ishizaka, Masamichi; Horisawa, Kenichi; Tateyama, Seiji; Takashima, Hideaki; Fuse, Shinichiro; Sue, Kaori; Hirai, Naoya; Masuoka, Kazuyo; Yanagawa, Hiroshi.

:: Genome Research, 巻 15, 番号 5, 01.05.2005, p. 710-717.

研究成果: ジャーナルへの寄稿記事

Miyamoto-Sato, E, Ishizaka, M, Horisawa, K, Tateyama, S, Takashima, H, Fuse, S, Sue, K, Hirai, N, Masuoka, K & Yanagawa, H 2005, 'Cell-free cotranslation and selection using in vitro virus for high-throughput analysis of protein-protein interactions and complexes', Genome Research, 巻. 15, 番号 5, pp. 710-717. https://doi.org/10.1101/gr.3510505
Miyamoto-Sato, Etsuko ; Ishizaka, Masamichi ; Horisawa, Kenichi ; Tateyama, Seiji ; Takashima, Hideaki ; Fuse, Shinichiro ; Sue, Kaori ; Hirai, Naoya ; Masuoka, Kazuyo ; Yanagawa, Hiroshi. / Cell-free cotranslation and selection using in vitro virus for high-throughput analysis of protein-protein interactions and complexes. :: Genome Research. 2005 ; 巻 15, 番号 5. pp. 710-717.
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AU - Fuse, Shinichiro

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