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Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae

  • Applied microbial and cell physiology
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Abstract

Monoterpenes have wide applications in the food, cosmetics, and medicine industries and have recently received increased attention as advanced biofuels. However, compared with sesquiterpenes, monoterpene production is still lagging in Saccharomyces cerevisiae. In this study, geraniol, a valuable acyclic monoterpene alcohol, was synthesized in S. cerevisiae. We evaluated three geraniol synthases in S. cerevisiae, and the geraniol synthase Valeriana officinalis (tVoGES), which lacked a plastid-targeting peptide, yielded the highest geraniol production. To improve geraniol production, synthesis of the precursor geranyl diphosphate (GPP) was regulated by comparing three specific GPP synthase genes derived from different plants and the endogenous farnesyl diphosphate synthase gene variants ERG20 G (ERG20 K197G) and ERG20 WW (ERG20 F96W-N127W), and controlling endogenous ERG20 expression, coupled with increasing the expression of the mevalonate pathway by co-overexpressing IDI1, tHMG1, and UPC2-1. The results showed that overexpressing ERG20 WW and strengthening the mevalonate pathway significantly improved geraniol production, while expressing heterologous GPP synthase genes or down-regulating endogenous ERG20 expression did not show positive effect. In addition, we constructed an Erg20p(F96W-N127W)-tVoGES fusion protein, and geraniol production reached 66.2 mg/L after optimizing the amino acid linker and the order of the proteins. The best strain yielded 293 mg/L geraniol in a fed-batch cultivation, a sevenfold improvement over the highest titer previously reported in an engineered S. cerevisiae strain. Finally, we showed that the toxicity of geraniol limited its production. The platform developed here can be readily used to synthesize other monoterpenes.

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Acknowledgments

This work was supported by the National High Technology Research and Development Program of China (2012AA022106), the National Natural Science Foundation of China (31470163), the National Key Technology R&D Program of China (2014BAD02B07), Project of the National Energy Administration of China (NY20130402), and the Key R & D Program of Shandong Province (2015GSF121015). We thank Prof. Hal S Alper for supplying the plasmid containing the UASCIT(3×) -TEF1 promoter.

Author contributions

JZ and JH designed the experiments. JZ and CL carried out the experiments. JZ, XB, YS, and JH analyzed the data. JZ and JH wrote the manuscript. All of the authors have read and approved the final manuscript.

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Authors and Affiliations

  1. State Key Laboratory of Microbial Technology, Shandong University, Jinan, 250100, China

    Jianzhi Zhao, Xiaoming Bao, Chen Li, Yu Shen & Jin Hou

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  1. Jianzhi Zhao
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  2. Xiaoming Bao
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  3. Chen Li
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  4. Yu Shen
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  5. Jin Hou
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Correspondence to Jin Hou.

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Jianzhi Zhao and Xiaoming Bao contributed equally to the study.

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Zhao, J., Bao, X., Li, C. et al. Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae . Appl Microbiol Biotechnol 100, 4561–4571 (2016). https://doi.org/10.1007/s00253-016-7375-1

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