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Abstract
BackgroundAlpha-Terpineol (α-Terpineol), a C10 monoterpenoid alcohol, is widely used in the cosmetic and pharmaceutical industries. Construction Saccharomyces cerevisiae cell factories for producing monoterpenes offers a promising means to substitute chemical synthesis or phytoextraction.Resultsα-Terpineol was produced by expressing the truncated α-Terpineol synthase (tVvTS) from Vitis vinifera in S. cerevisiae. The α-Terpineol titer was increased to 0.83 mg/L with overexpression of the rate-limiting genes tHMG1, IDI1 and ERG20F96W-N127W. A GSGSGSGSGS linker was applied to fuse ERG20F96W-N127W with tVvTS, and expressing the fusion protein increased the α-Terpineol production by 2.87-fold to 2.39 mg/L when compared with the parental strain. In addition, we found that farnesyl diphosphate (FPP) accumulation by down-regulation of ERG9 expression and deletion of LPP1 and DPP1 did not improve α-Terpineol production. Therefore, ERG9 was overexpressed and the α-Terpineol titer was further increased to 3.32 mg/L. The best α-Terpineol producing strain LCB08 was then used for batch and fed-batch fermentation in a 5 L bioreactor, and the production of α-Terpineol was ultimately improved to 21.88 mg/L.ConclusionsAn efficient α-Terpineol production cell factory was constructed by engineering the S. cerevisiae mevalonate pathway, and the metabolic engineering strategies could also be applied to produce other valuable monoterpene compounds in yeast.
Highlights
Alpha-Terpineol (α-Terpineol), a C10 monoterpenoid alcohol, is widely used in the cosmetic and pharmaceutical industries
Isopentenyl diphosphate (IPP) and dimethylallyl pyrophosphate (DMAPP) are the universal C5 precursors for synthesizing monoterpenoids (C10), sesquiterpenoids (C15), diterpenoids (C20) and triterpenoids (C30) among other terpenoid compounds [2]. α-Terpineol with the chemical formula C10H180 is a monoterpenoid compound existing in plants; it was often used as perfume and repellent in the cosmetic industry and as an anticonvulsant agent in the pharmaceutical industry [3, 4]. α-Terpineol can be used
MAF1, a negative regulator of tRNA biosynthesis that shares the same precursors as geranyl diphosphate (GPP), was reported to have positive effects on geraniol production when deleted in S. cerevisiae [18]
Summary
Alpha-Terpineol (α-Terpineol), a C10 monoterpenoid alcohol, is widely used in the cosmetic and pharmaceutical industries. The engineered GPP synthase was expressed in diploid S. cerevisiae cells, and the resulting strain displayed a significant increase in monoterpene sabinene yield [17]. MAF1, a negative regulator of tRNA biosynthesis that shares the same precursors as GPP, was reported to have positive effects on geraniol production when deleted in S. cerevisiae [18]. A CEN.PK-113-5D S. cerevisiae strain was engineered to produce monoterpene linalool and yielded a two-fold titer increase with ERG9 down-regulation [19]. These completely contradictory results prompted us to study the influence of the downstream MVA pathway on monoterpene synthesis in S. cerevisiae
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