Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2025 Jun 23:13:1621955.
doi: 10.3389/fbioe.2025.1621955. eCollection 2025.

Enabling and improving trans-nerolidol production by Corynebacterium glutamicum: combining metabolic engineering and trace elements medium refinement

Jan Seeger  1 , Stella Lohoff  1 , Fabian Schmitfranz  1 , Nadja A Henke  1 , Volker F Wendisch  1
Affiliations

Enabling and improving trans-nerolidol production by Corynebacterium glutamicum: combining metabolic engineering and trace elements medium refinement

Jan Seeger et al. Front Bioeng Biotechnol. .

Abstract

Terpenes are biomolecules of significant industrial relevance, with applications in pharmaceuticals, cosmetics, and the food industry. Their biotechnological production is emerging, with Corynebacterium glutamicum, a Gram-positive bacterium traditionally employed for large-scale amino acid production, serving as a promising host. While metabolic engineering strategies have been extensively applied to enhance terpene titers in C. glutamicum, the role of medium composition, particularly trace elements, remains underexplored. In this study, the impact of trace element composition on trans-nerolidol production by engineered C. glutamicum was investigated. A Design of Experiments (DoE) approach identified MgSO4 as a critical factor, and the refined trace element composition led to a 34% increase in trans-nerolidol production. Further metabolic engineering efforts resulted in a final titer of 28.1 mg L-1. Subsequent fed-batch fermentation achieved a trans-nerolidol titer of 0.41 g L-1, representing the highest reported sesquiterpene titer being produced by C. glutamicum to date. Additionally, the refined trace element composition was successfully applied to patchoulol- and (+)-valencene-producing strains, leading to production increases of 15% and 72%, respectively. These findings demonstrate that trace element refinement and metabolic engineering act as complementary strategies for enhancing terpene production in a microbial production host.

Keywords: Corynebacterium glutamicum; design of experiment; media optimization; metabolic engineering; terpenes; trans-nerolidol.

PubMed Disclaimer

Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Biosynthetic pathway for trans-nerolidol production in Corynebacterium glutamicum. Dashed arrows represent multiple enzymatic steps, gene deletions are marked with red crosses, Mg2+ dependency of key enzymes is indicated in blue. Gene overexpression is indicated by green plus symbols; the number of symbols refers to expression strength. Abbreviations: GAP, glyceraldehyde 3-phosphate; DXP, 1-deoxy-D-xylulose 5-phosphate; IPP, isopentenyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; FPP, farnesyl pyrophosphate; GGPP, geranylgeranyl pyrophosphate; DXS, DXP synthase; MEP, 2-C-methyl-D-erythritol 4-phosphate; Idi, IPP isomerase; IspAEc, FPP synthase from Escherichia coli; NSTw, Nerolidol synthase from Tripterygium wilfordii; IdsA, GGPP synthase; CrtE, GGPP synthase.
FIGURE 2
FIGURE 2
trans-Nerolidol production by C. glutamicum NERO1 and NERO2. Means, standard deviations as error bars, and single points of triplicate cultivations are given. Statistical significance was calculated with a Students’ t-test p < 0.05 (*), p < 0.01 (**), p < 0.005 (***).
FIGURE 3
FIGURE 3
Steepest ascent experiment based on the first CCD. Means, standard deviations as error bars, and single points of triplicate cultivations are given. Statistical significance was calculated with a Students’ t-test p < 0.05 (*), p < 0.01 (**), p < 0.005 (***).
FIGURE 4
FIGURE 4
Comparison of different trace element compositions using strain NERO2. From left to right: standard CGXII trace elements, best composition of steepest ascent experiment, standard CGXII trace elements with 4x MgSO4 concentration, best composition of second CCD experiment (=refined). Means, standard deviations as error bars, and single points of triplicate cultivations are given. Statistical significance was calculated with a Students’ t-test p < 0.05 (*), p < 0.01 (**), p < 0.005 (***).
FIGURE 5
FIGURE 5
Comparison of different trans-nerolidol producing strains with standard and refined trace elements. Means, standard deviations as error bars, and single points of triplicate cultivations are given. Statistical significance was calculated with a Students’ t-test p < 0.05 (*), p < 0.01 (**), p < 0.005 (***).
FIGURE 6
FIGURE 6
Fed-batch fermentation of NERO5. Cell dry weight concentration (CDW) and trans-nerolidol titer are given.
FIGURE 7
FIGURE 7
Comparison of standard CGXII with refined trace elements for the production of (+)-valencene and patchoulol. Means, standard deviations as error bars, and single points of triplicate cultivations are given. Statistical significance was calculated with a Students’ t-test p < 0.05 (*), p < 0.01 (**), p < 0.005 (***).
See this image and copyright information in PMC

Similar articles

See all similar articles

References

    1. Adamczyk P. A., Hwang H. J., Chang T.-H., Gao Y., Baidoo E. E. K., Kim J., et al. (2025). The oleaginous yeast Rhodosporidium toruloides engineered for biomass hydrolysate-derived (E)-α-bisabolene production. Metab. Eng. 90, 92–105. 10.1016/j.ymben.2025.02.014 - DOI - PubMed
    1. Ambrož M., Boušová I., Skarka A., Hanušová V., Králová V., Matoušková P., et al. (2015). The influence of sesquiterpenes from Myrica rubra on the antiproliferative and pro-oxidative effects of doxorubicin and its accumulation in cancer cells. Molecules 20, 15343–15358. 10.3390/molecules200815343 - DOI - PMC - PubMed
    1. Bertani G. (1951). Studies on lysogenesis I. J. Bacteriol. 62, 293–300. 10.1128/jb.62.3.293-300.1951 - DOI - PMC - PubMed
    1. Binder D., Frohwitter J., Mahr R., Bier C., Grünberger A., Loeschcke A., et al. (2016). Light-controlled cell factories: employing photocaged isopropyl-β-D-thiogalactopyranoside for light-mediated optimization of lac promoter-based gene expression and (+)-valencene biosynthesis in Corynebacterium glutamicum . Appl. Environ. Microbiol. 82, 6141–6149. 10.1128/aem.01457-16 - DOI - PMC - PubMed
    1. Buchholz J., Graf M., Blombach B., Takors R. (2014). Improving the carbon balance of fermentations by total carbon analyses. Biochem. Eng. J. 90, 162–169. 10.1016/J.BEJ.2014.06.007 - DOI

LinkOut - more resources