A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption

Maiko Furubayashi, Mayu Ikezumi, Jun Kajiwara, Miki Iwasaki, Akira Fujii, Ng Li, Kyoichi Saito, Daisuke Umeno

研究成果: Article査読

32 被引用数 (Scopus)

抄録

Terpene synthases catalyze the formation of a variety of terpene chemical structures. Systematic mutagenesis studies have been effective in providing insights into the characteristic and complex mechanisms of C-C bond formations and in exploring the enzymatic potential for inventing new chemical structures. In addition, there is growing demand to increase terpene synthase activity in heterologous hosts, given the maturation of metabolic engineering and host breeding for terpenoid synthesis. We have developed a simple screening method for the cellular activities of terpene synthases by scoring their substrate consumption based on the color loss of the cell harboring carotenoid pathways. We demonstrate that this method can be used to detect activities of various terpene synthase or prenyltransferase genes in a high-throughput manner, irrespective of the product type, enabling the mutation analysis and directed evolution of terpene synthases. We also report the possibility for substrate-specific screening system of terpene synthases by taking advantage of the substrate-size specificity of C30 and C40 carotenoid pathways.

本文言語English
論文番号e93317
ジャーナルPloS one
9
3
DOI
出版ステータスPublished - 2014 3 28
外部発表はい

ASJC Scopus subject areas

  • 生化学、遺伝学、分子生物学(全般)
  • 農業および生物科学(全般)
  • 一般

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