Increased carvone production in Escherichia coli by balancing limonene conversion enzyme expression via targeted quantification concatamer proteome analysis
Abstract (−)-Carvone is a monoterpenoid with a spearmint flavor. A sustainable biotechnological production process for (−)-carvone is desirable. Although all enzymes in (−)-carvone biosynthesis have been functionally expressed in Escherichia coli independently, the yield was low in previous studies....
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| Autores principales: | , , , , , , , , , |
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| Formato: | article |
| Lenguaje: | EN |
| Publicado: |
Nature Portfolio
2021
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| Materias: | |
| Acceso en línea: | https://doaj.org/article/5e010c658d71455cb769685da713f645 |
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| Sumario: | Abstract (−)-Carvone is a monoterpenoid with a spearmint flavor. A sustainable biotechnological production process for (−)-carvone is desirable. Although all enzymes in (−)-carvone biosynthesis have been functionally expressed in Escherichia coli independently, the yield was low in previous studies. When cytochrome P450 limonene-6-hydroxylase (P450)/cytochrome P450 reductase (CPR) and carveol dehydrogenase (CDH) were expressed in a single strain, by-product formation (dihydrocarveol and dihydrocarvone) was detected. We hypothesized that P450 and CDH expression levels differ in E. coli. Thus, two strains independently expressing P450/CPR and CDH were mixed with different ratios, confirming increased carvone production and decreased by-product formation when CDH input was reduced. The optimum ratio of enzyme expression to maximize (−)-carvone production was determined using the proteome analysis quantification concatamer (QconCAT) method. Thereafter, a single strain expressing both P450/CPR and CDH was constructed to imitate the optimum expression ratio. The upgraded strain showed a 15-fold improvement compared to the initial strain, showing a 44 ± 6.3 mg/L (−)-carvone production from 100 mg/L (−)-limonene. Our study showed the usefulness of the QconCAT proteome analysis method for strain development in the industrial biotechnology field. |
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