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A bacterial platform for producing aromatic esters from glycerol

Nature Chemical Engineering volume 1, pages 751–764 (2024)Cite this article

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Abstract

Aromatic esters possess flavor and fragrance qualities that are widely used in the food, pharmaceutical and cosmetic industries. However, microbial production of these compounds is hampered by a limited understanding of the natural biosynthetic pathway and the relatively low titer and yield. This study establishes a microbial platform for the efficient production of various aromatic esters. A systematic engineering strategy was developed, involving reshaping the substrate access tunnel to enhance enzyme substrate specificity, shifting acetyl coenzyme A metabolic pathways to improve cofactor supply and engineering a dynamic regulation system to redistribute the carbon flux from cell growth toward product synthesis. The implementation of these approaches led to the production of 10.4 g l–1 benzyl benzoate, representing a 4,700-fold increase in titer compared with the initial strain. This work showcases a bacterial platform for the efficient production of aromatic esters and offers insights into overcoming challenges in microbial cell factory construction.

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Fig. 1: Designing and assembling an artificial biosynthetic pathway for BBZ production.
Fig. 2: Enhancing the supply of BA to achieve de novo biosynthesis of BBZ.
Fig. 3: Reshaping the substrate access tunnel of Car to prevent by-product formation.
Fig. 4: Improving BBZ titer by boosting cofactor availability.
Fig. 5: Increasing the BBZ titer by dynamically controlling gltA* expression.
Fig. 6: Extending the bacterial platform for biosynthesis of various aromatic esters.

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Data availability

The data supporting the results are provided in the article and its Supplementary Information. Related materials, including the protein structures, tunnel analyses, docking simulations, ASMD simulations, MD simulations, CAVER Dock analyses and a video showing the dynamic changes of all four tunnels throughout the MD simulations are available via Zenodo at https://doi.org/10.5281/zenodo.12784348 (ref. 78). Source data are provided with this paper.

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Acknowledgements

This work was supported by the National Key Research and Development Program of China (grant no. 2022YFC2106100) and the National Natural Science Foundation of China (grant nos. 22378016, 22078011 and 22238001). We thank Z. Sun and C. Li (Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences) for their guidance on the ASMD simulation and Y. Zhou (Dalian Institute of Chemical Physics, Chinese Academy of Sciences) for his guidance on paper preparation.

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Authors and Affiliations

  1. State Key Laboratory of Chemical Resource Engineering, Beijing University of Chemical Technology, Beijing, China

    Liangyu Lu, Xiaolei Wang, Tong Wang, Xiaolin Shen, Xinxiao Sun, Pingfang Tian, Jia Wang & Qipeng Yuan

  2. Beijing Advanced Innovation Center for Soft Matter Science and Engineering, Beijing University of Chemical Technology, Beijing, China

    Liangyu Lu, Xiaolin Shen, Xinxiao Sun, Pingfang Tian, Jens Nielsen, Jia Wang & Qipeng Yuan

  3. School of Chemical, Materials, and Biomedical Engineering, College of Engineering, The University of Georgia, Athens, GA, USA

    Yajun Yan

  4. Department of Life Sciences, Chalmers University of Technology, Gothenburg, Sweden

    Jens Nielsen

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  1. Liangyu Lu
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Contributions

L.L. and J.W. conceived the study and wrote the paper. L.L., X.W. and T.W. performed the experiments. X. Shen and X. Sun participated in the research. J.W., P.T., Y.Y., J.N. and Q.Y. revised the paper. J.W. and Q.Y. directed the research. All authors contributed to the article and approved the submitted version.

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Correspondence to Jia Wang or Qipeng Yuan.

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Nature Chemical Engineering thanks Guo-Qiang Chen, Yong Hwan Kim, Cong T. Trinh and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Extended data

Extended Data Table 1 Kinetic parameters of the wildtype Car and its variants toward CA and BA
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Supplementary Information

Supplementary Figs. 1–21, Tables 1–3 and references.

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Supplementary Data 1

Oligonucleotides used in this study.

Supplementary Data 2

Nonsense sites in the E. coli genome and corresponding sgRNA (N20).

Supplementary Data 3

Raw NMR data.

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Lu, L., Wang, X., Wang, T. et al. A bacterial platform for producing aromatic esters from glycerol. Nat Chem Eng 1, 751–764 (2024). https://doi.org/10.1038/s44286-024-00148-9

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