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Engineering living cells with polymers for recyclable photoenzymatic catalysis

Nature Catalysis volume 7, pages 1404–1416 (2024)Cite this article

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Abstract

Engineering cell membranes for catalysis is challenging due to their inherent complexity. Here we introduce a polymeric strategy to overcome these challenges by chemically modifying cell membranes with catalytic polymers, enabling robust, recyclable and photoenzymatic catalysis. Through a one-step in situ atom transfer radical polymerization on living Escherichia coli cells, polymers are generated to protect the cells from environmental stressors while facilitating chemoenzymatic synthesis by integrating catalytic polymers with intracellular enzymes. As a proof of concept, a photoenzymatic cascade with an anthraquinone-based polymer and benzaldehyde lyase is demonstrated, converting benzyl alcohol into benzoin and achieving bioconversion yields that are 15 times higher than controls. Additionally, cells serve as large biological scaffolds for polymers, enabling recycling of macromolecular catalysts. A recyclable chemoenzymatic system incorporating an organometallic polymer with intracellular enzymes is also presented. Our versatile, straightforward approach offers a technology platform for engineering cell membranes for cascade synthesis, with broad implications for synthetic chemistry, polymer chemistry and biotechnology.

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Fig. 1: Fabrication of polymer-grafted cells and their applications in chemoenzymatic cascades.
Fig. 2: Characterization of grafted polymers and their distribution on cell surfaces.
Fig. 3: Viability and proliferation investigations of polymer-grafted E. coli cells.
Fig. 4: Compatibility evaluation of photo- and biocatalysts in polymer-grafted E. coli cells.
Fig. 5: Polymeric protection against external stress environment.
Fig. 6: Photoenzymatic cascade of polymer-grafted E. coli cells.
Fig. 7: Chemoenzymatic cascade studies of polymer-grafted E. coli cells.

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Data availability

All data supporting the findings of this study are available in the Article and the Supplementary Information or are available from the corresponding author upon reasonable request. Source data are provided with this paper.

References

  1. Hothorn, M. et al. Catalytic core of a membrane-associated eukaryotic polyphosphate polymerase. Science 324, 513–516 (2009).

    CAS  PubMed  Google Scholar 

  2. Suladze, S., Cinar, S., Sperlich, B. & Winter, R. Pressure modulation of the enzymatic activity of phospholipase A2, a putative membrane-associated pressure sensor. J. Am. Chem. Soc. 137, 12588–12596 (2015).

    CAS  PubMed  Google Scholar 

  3. Hendrickx, A., Budzik, J., Young, S. & Schneewind, O. Architects at the bacterial surface—sortases and the assembly of pili with isopeptide bonds. Nat. Rev. Microbiol. 9, 166–176 (2011).

    CAS  PubMed  Google Scholar 

  4. Hussain, M. S., Wang, Q. Z. & Viola, R. E. Engineering of a critical membrane-anchored enzyme for high solubility and catalytic activity. Arch. Biochem. Biophys. 703, 108870 (2021).

    CAS  PubMed  Google Scholar 

  5. Mu, J. et al. A small-molecule FRET reporter for the real-time visualization of cell-surface proteolytic enzyme functions. Angew. Chem. Int. Ed. 53, 14357–14362 (2014).

    CAS  Google Scholar 

  6. Thomas, G. Furin at the cutting edge: from protein traffic to embryogenesis and disease. Nat. Rev. Mol. Cell Biol. 3, 753–766 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Seidah, N. G. & Prat, A. The biology and therapeutic targeting of the proprotein convertases. Nat. Rev. Drug Discov. 11, 367–383 (2012).

    CAS  PubMed  Google Scholar 

  8. Lai, X., Wichers, H. J., S, L. M. & Dijkstra, B. W. Structure of human tyrosinase related protein 1 reveals a binuclear zinc active site important for melanogenesis. Angew. Chem. Int. Ed. 56, 9812–9815 (2017).

    CAS  Google Scholar 

  9. Grodd, W., Krägeloh-Mann, I., Petersen, D., Trefz, F. K. & Harzer, K. In vivo assessment of N-acetylaspartate in brain in spongy degeneration (Canavan’s disease) by proton spectroscopy. Lancet 336, 437–438 (1990).

    CAS  PubMed  Google Scholar 

  10. Bohl, H. O., Shi, K., Lee, J. K. & Aihara, H. Crystal structure of lipid A disaccharide synthase LpxB from Escherichia coli. Nat. Commun. 9, 377 (2018).

    PubMed  PubMed Central  Google Scholar 

  11. Gómez, M. S. et al. The oxidative fermentation of ethanol in Gluconacetobacter diazotrophicus is a two-step pathway catalyzed by a single enzyme: alcohol-aldehyde dehydrogenase (ADHa). Int. J. Mol. Sci. 16, 1293–1311 (2015).

    Google Scholar 

  12. Yakushi, T. & Matsushita, K. Alcohol dehydrogenase of acetic acid bacteria: structure, mode of action, and applications in biotechnology. Appl. Microbiol. Biotechnol. 86, 1257–1265 (2010).

    CAS  PubMed  Google Scholar 

  13. Yang, H. et al. Molecular biology: fantastic toolkits to improve knowledge and application of acetic acid bacteria. Biotechnol. Adv. 58, 107911 (2022).

    CAS  PubMed  Google Scholar 

  14. Fan, L. H., Zhang, Z. J., Yu, X. Y., Xue, Y. X. & Tan, T. W. Self-surface assembly of cellulosomes with two miniscaffoldins on Saccharomyces cerevisiae for cellulosic ethanol production. Proc. Natl Acad. Sci. USA 109, 13260–13265 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Tanaka, T. & Kondo, A. Cell surface engineering of industrial microorganisms for biorefining applications. Biotechnol. Adv. 33, 1403–1411 (2015).

    CAS  PubMed  Google Scholar 

  16. Chen, K. & Arnold, F. H. Engineering new catalytic activities in enzymes. Nat. Catal. 3, 203–213 (2020).

    CAS  Google Scholar 

  17. Lee, S. Y., Choi, J. H. & Xu, Z. Microbial cell-surface display. Trends Biotechnol. 21, 45–52 (2003).

    CAS  PubMed  Google Scholar 

  18. Smith, G. P. Filamentous fusion phage: novel expression vectors that display cloned antigens on the virion surface. Science 228, 1315–1317 (1985).

    CAS  PubMed  Google Scholar 

  19. Charbit, A., Boulain, J. C., Ryter, A. & Hofnung, M. Probing the topology of a bacterial membrane protein by genetic insertion of a foreign epitope; expression at the cell surface. EMBO J. 5, 3029–3037 (1986).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Richins, R. D., Kaneva, I., Mulchandani, A. & Chen, W. Biodegradation of organophosphorus pesticides by surface-expressed organophosphorus hydrolase. Nat. Biotechnol. 15, 984–987 (1997).

    CAS  PubMed  Google Scholar 

  21. Jin, Z. et al. Combined utilization of lipase-displaying Pichia pastoris whole-cell biocatalysts to improve biodiesel production in co-solvent media. Bioresour. Technol. 130, 102–109 (2013).

    CAS  PubMed  Google Scholar 

  22. Boder, E. T. & Wittrup, K. D. Yeast surface display for screening combinatorial polypeptide libraries. Nat. Biotechnol. 15, 553–557 (1997).

    CAS  PubMed  Google Scholar 

  23. Szponarski, M., Schwizer, F., Ward, T. R. & Gademann, K. On-cell catalysis by surface engineering of live cells with an artificial metalloenzyme. Commun. Chem. 1, 84 (2018).

    Google Scholar 

  24. Schmid, A. et al. Industrial biocatalysis today and tomorrow. Nature 409, 258–268 (2001).

    CAS  PubMed  Google Scholar 

  25. Stepankova, V. et al. Strategies for stabilization of enzymes in organic solvents. ACS Catal. 3, 2823–2836 (2013).

    CAS  Google Scholar 

  26. Zhang, N., Sun, Z. & Wu, C. Artificial enzymes combining proteins with proline polymers for asymmetric aldol reactions in water. ACS Catal. 12, 4777–4783 (2022).

    CAS  Google Scholar 

  27. Chi, Y., Scroggins, S. T., Boz, E. & Fréchet, J. M. J. Control of aldol reaction pathways of enolizable aldehydes in an aqueous environment with a hyperbranched polymeric catalyst. J. Am. Chem. Soc. 130, 17287–17289 (2008).

    CAS  PubMed  Google Scholar 

  28. Liu, F., Kong, W., Qi, C., Zhu, L. & Xiao, F. S. Design and synthesis of mesoporous polymer-based solid acid catalysts with excellent hydrophobicity and extraordinary catalytic activity. ACS Catal. 2, 565–572 (2012).

    CAS  Google Scholar 

  29. Huang, K., Zhang, J. Y., Liu, F. & Dai, S. Synthesis of porous polymeric catalysts for the conversion of carbon dioxide. ACS Catal. 8, 9079–9102 (2018).

    CAS  Google Scholar 

  30. Sun, Z., Hübner, R., Li, J. & Wu, C. Artificially sporulated Escherichia coli cells as a robust cell factory for interfacial biocatalysis. Nat. Commun. 13, 3142 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Park, J. H. et al. A cytoprotective and degradable metal–polyphenol nanoshell for single-cell encapsulation. Angew. Chem. Int. Ed. 53, 12420–12425 (2014).

    CAS  Google Scholar 

  32. Kim, J. Y. et al. Cytocompatible polymer grafting from individual living cells by atom-transfer radical polymerization. Angew. Chem. Int. Ed. 55, 15306–15309 (2016).

    CAS  Google Scholar 

  33. Magennis, E. P. et al. Bacteria-instructed synthesis of polymers for self-selective microbial binding and labelling. Nat. Mater. 13, 748–755 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Niu, J. et al. Engineering live cell surfaces with functional polymers via cytocompatible controlled radical polymerization. Nat. Chem. 9, 537–545 (2017).

    CAS  PubMed  Google Scholar 

  35. Song, R. B. et al. Living and conducting: coating individual bacterial cells with in situ formed polypyrrole. Angew. Chem. Int. Ed. 56, 10516–10520 (2017).

    CAS  Google Scholar 

  36. Laskar, P., Varghese, O. P. & Shastri, V. P. Advances in intracellular and on-surface polymerization in living cells: implications for nanobiomedicines. Adv. Nanobiomed. Res. 3, 2200174 (2023).

    CAS  Google Scholar 

  37. Pan, J. et al. A single-cell nanocoating of probiotics for enhanced amelioration of antibiotic-associated diarrhea. Nat. Commun. 13, 2117 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Smith, P. N., Mao, L., Sinha, K. & Russell, A. J. Organophosphate detoxification by membrane-engineered red blood cells. Acta Biomater. 124, 270–281 (2021).

    CAS  PubMed  Google Scholar 

  39. Safarova, Y. et al. Mesenchymal stem cells coated with synthetic bone-targeting polymers enhance osteoporotic bone fracture regeneration. Bioengineering 7, 125 (2020).

    CAS  Google Scholar 

  40. Clafshenkel, W. P. et al. The effect of covalently-attached ATRP-synthesized polymers on membrane stability and cytoprotection in human erythrocytes. PLoS One 11, e0157641 (2016).

    PubMed  PubMed Central  Google Scholar 

  41. D’Souza, S. et al. Engineering of cell membranes with a bisphosphonate-containing polymer using ATRP synthesis for bone targeting. Biomaterials 35, 9447–9458 (2014).

    PubMed  Google Scholar 

  42. Teramura, Y. & Iwata, H. Cell surface modification with polymers for biomedical studies. Soft Matter 6, 1081–1091 (2010).

    CAS  Google Scholar 

  43. Teramura, Y., Kaneda, Y. & Iwata, H. Islet-encapsulation in ultra-thin layer-by-layer membranes of poly(vinyl alcohol) anchored to poly(ethylene glycol)-lipids in the cell membrane. Biomaterials 28, 4818–4825 (2007).

    CAS  PubMed  Google Scholar 

  44. Sun, Z., Glebe, U., Charan, H., Böker, A. & Wu, C. Enzyme–polymer conjugates as robust pickering interfacial biocatalysts for efficient biotransformations and one-pot cascade reactions. Angew. Chem. Int. Ed. 57, 13810–13814 (2018).

    CAS  Google Scholar 

  45. Wang, J. S. & Matyjaszewski, K. Controlled/‘living’ radical polymerization. atom transfer radical polymerization in the presence of transition-metal complexes. J. Am. Chem. Soc. 117, 5614–5615 (1995).

    CAS  Google Scholar 

  46. Matyjaszewski, K. & Tsarevsky, N. V. Macromolecular engineering by atom transfer radical polymerization. J. Am. Chem. Soc. 136, 6513–6533 (2014).

    CAS  PubMed  Google Scholar 

  47. Vargason, A. M., Santhosh, S. & Anselmo, A. C. Surface modifications for improved delivery and function of therapeutic bacteria. Small 16, 2001705 (2020).

    CAS  Google Scholar 

  48. Li, D. et al. Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics. Science 349, aab3500 (2015).

    PubMed  PubMed Central  Google Scholar 

  49. Sahl, S. J., Hell, S. W. & Jakobs, S. Fluorescence nanoscopy in cell biology. Nat. Rev. Mol. Cell Biol. 18, 685–701 (2017).

    CAS  PubMed  Google Scholar 

  50. Kraus, F. et al. Quantitative 3D structured illumination microscopy of nuclear structures. Nat. Protoc. 12, 1011–1028 (2017).

    CAS  PubMed  Google Scholar 

  51. Boulos, L., Prévost, M., Barbeau, B., Coallier, J. & Desjardins, R. LIVE/DEAD BacLight: application of a new rapid staining method for direct enumeration of viable and total bacteria in drinking water. J. Microbiol. Methods 37, 77–86 (1999).

    CAS  PubMed  Google Scholar 

  52. Köhler, V. et al. Synthetic cascades are enabled by combining biocatalysts with artificial metalloenzymes. Nat. Chem. 5, 93–99 (2013).

    PubMed  Google Scholar 

  53. Zhang, W., Gacs, J., Isabel, W. C. E. & Hollmann, F. Selective photooxidation reactions using water-soluble anthraquinone photocatalysts. ChemCatChem 9, 3821–3826 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  54. Imlay, J. A. The molecular mechanisms and physiological consequences of oxidative stress: lessons from a model bacterium. Nat. Rev. Microbiol. 11, 443–454 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We are grateful for financial support from the Independent Research Fund Denmark within the framework of the Sapere Aude leader programme (9064-00062B, C.W.). We also thank the Novo Nordisk Foundation (NNF21OC0071661, C.W.), the Carlsberg Foundation (CF22-1008, C.W.), the S.C. VAN foundation (J.N.) and the National Key R&D Program of China (2023YFA0913600, Z.S.) for their generous funding. We are grateful to Enzymicals AG for providing the plasmid of ATA, to A. Worbs for performing SEM and to C. He (Sichuan University) for TEM and EDXS measurements. We would like to acknowledge the assistance of the core facility BioSupraMol (FU Berlin), C. Hudziak (FU Berlin) for GPC measurement and T. Frickmann and D. E. Canfield (University of Southern Denmark) for ICP-MS analysis. The use of the HZDR Ion Beam Center TEM facilities is acknowledged. Cryo-TEM image acquisition and image analyses were performed at the Institute of Chemistry and Biochemistry, Electron Microscopy Research Center (FU Berlin). Fluorescence image acquisition and image analyses were performed at the Danish Molecular Biomedical Imaging Center (DaMBIC, University of Southern Denmark), supported by the Novo Nordisk Foundation (NNF18SA0032928, M.F.E.).

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Author notes

  1. These authors contributed equally: Jian Ning, Zhiyong Sun.

Authors and Affiliations

  1. Department of Physics, Chemistry and Pharmacy, University of Southern Denmark, Odense, Denmark

    Jian Ning & Changzhu Wu

  2. College of Biotechnology and Bioengineering, Zhejiang University of Technology, Hangzhou, China

    Zhiyong Sun

  3. Helmholtz-Zentrum Dresden-Rossendorf, Institute of Ion Beam Physics and Materials Research, Dresden, Germany

    René Hübner

  4. Department of Green Technology, University of Southern Denmark, Odense, Denmark

    Henrik Karring

  5. DaMBIC, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense, Denmark

    Morten Frendø Ebbesen

  6. Department of Biology, Chemistry and Pharmacy, Institute of Chemistry and Biochemistry, Research Center of Electron Microscopy (FZEM), Free University of Berlin, Berlin, Germany

    Mathias Dimde

  7. Danish Institute for Advanced Study (DIAS), University of Southern Denmark, Odense, Denmark

    Changzhu Wu

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Contributions

C.W. conceptualized and supervised the project. J.N. performed the experiments. Z.S. provided experimental guidance and scientific discussion. R.H. performed SEM-based data analysis. M.F.E. conducted SIM and STED measurements. M.D. performed cryo-TEM and relevant data analysis. H.K. provided guidance and supervision related to the microbiological work. C.W., J.N. and Z.S. wrote the paper.

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Correspondence to Changzhu Wu.

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Nature Catalysis thanks Nico Bruns, Florian Rudroff and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Supplementary methods, Table 1, Figs. 1–37 and Notes 1 and 2.

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Ning, J., Sun, Z., Hübner, R. et al. Engineering living cells with polymers for recyclable photoenzymatic catalysis. Nat Catal 7, 1404–1416 (2024). https://doi.org/10.1038/s41929-024-01259-5

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