Identification of the sesquiterpene synthase AcTPS1 and high production of (-)-germacrene D in metabolically engineered Saccharomyces cerevisiae.

Publisher: BioMed Central Country of Publication: England NLM ID: 101139812 Publication Model: Electronic Cited Medium: Internet ISSN: 1475-2859 (Electronic) Linking ISSN: 14752859 NLM ISO Abbreviation: Microb Cell Fact Subsets: MEDLINE

Original Publication: London : BioMed Central, [2002-

Saccharomyces cerevisiae*/genetics ; Saccharomyces cerevisiae*/metabolism ; Sesquiterpenes*/metabolism; Metabolic Engineering ; Sesquiterpenes, Germacrane/metabolism

Background: The sesquiterpene germacrene D is a highly promising product due to its wide variety of insecticidal activities and ability to serve as a precursor for many other sesquiterpenes. Biosynthesis of high value compounds through genome mining for synthases and metabolic engineering of microbial factories, especially Saccharomyces cerevisiae, has been proven to be an effective strategy. However, there have been no studies on the de novo synthesis of germacrene D from carbon sources in microbes. Hence, the construction of the S. cerevisiae cell factory to achieve high production of germacrene D is highly desirable.
Results: We identified five putative sesquiterpene synthases (AcTPS1 to AcTPS5) from Acremonium chrysogenum and the major product of AcTPS1 characterized by in vivo, in vitro reaction and NMR detection was revealed to be (-)-germacrene D. After systematically comparing twenty-one germacrene D synthases, AcTPS1 was found to generate the highest amount of (-)-germacrene D and was integrated into the terpene precursor-enhancing yeast strain, achieving 376.2 mg/L of (-)-germacrene D. Iterative engineering was performed to improve the production of (-)-germacrene D, including increasing the copy numbers of AcTPS1, tHMG1 and ERG20, and downregulating or knocking out other inhibitory factors (such as erg9, rox1, dpp1). Finally, the optimal strain LSc81 achieved 1.94 g/L (-)-germacrene D in shake-flask fermentation and 7.9 g/L (-)-germacrene D in a 5-L bioreactor, which is the highest reported (-)-germacrene D titer achieved to date.
Conclusion: We successfully achieved high production of (-)-germacrene D in S. cerevisiae through terpene synthase mining and metabolic engineering, providing an impressive example of microbial overproduction of high-value compounds.
(© 2022. The Author(s).)

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Keywords: Acremonium chrysogenum; Germacrene D; Metabolic engineering; Saccharomyces cerevisiae; Sesquiterpene

0 (Sesquiterpenes)
0 (Sesquiterpenes, Germacrane)
V2I9ATG34E (germacrene D)

Date Created: 20220518 Date Completed: 20220520 Latest Revision: 20220716

20231215

PMC9115970

10.1186/s12934-022-01814-4

35585553