A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption.
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 str...
| Main Authors: | , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2014-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC3969365?pdf=render |
| _version_ | 1818147418686881792 |
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| author | Maiko Furubayashi Mayu Ikezumi Jun Kajiwara Miki Iwasaki Akira Fujii Ling Li Kyoichi Saito Daisuke Umeno |
| author_facet | Maiko Furubayashi Mayu Ikezumi Jun Kajiwara Miki Iwasaki Akira Fujii Ling Li Kyoichi Saito Daisuke Umeno |
| author_sort | Maiko Furubayashi |
| collection | DOAJ |
| description | 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. |
| first_indexed | 2024-12-11T12:34:56Z |
| format | Article |
| id | doaj.art-67df3da6120549faabd3a1e971c4f049 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-11T12:34:56Z |
| publishDate | 2014-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-67df3da6120549faabd3a1e971c4f0492022-12-22T01:07:09ZengPublic Library of Science (PLoS)PLoS ONE1932-62032014-01-0193e9331710.1371/journal.pone.0093317A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption.Maiko FurubayashiMayu IkezumiJun KajiwaraMiki IwasakiAkira FujiiLing LiKyoichi SaitoDaisuke UmenoTerpene 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.http://europepmc.org/articles/PMC3969365?pdf=render |
| spellingShingle | Maiko Furubayashi Mayu Ikezumi Jun Kajiwara Miki Iwasaki Akira Fujii Ling Li Kyoichi Saito Daisuke Umeno A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. PLoS ONE |
| title | A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. |
| title_full | A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. |
| title_fullStr | A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. |
| title_full_unstemmed | A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. |
| title_short | A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. |
| title_sort | high throughput colorimetric screening assay for terpene synthase activity based on substrate consumption |
| url | http://europepmc.org/articles/PMC3969365?pdf=render |
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