Versatile biomanufacturing through stimulus-responsive cell–material feedback


Small-scale production of biologics has great potential for enhancing the accessibility of biomanufacturing. By exploiting cell–material feedback, we have designed a concise platform to achieve versatile production, analysis and purification of diverse proteins and protein complexes. The core of our technology is a microbial swarmbot, which consists of a stimulus-sensitive polymeric microcapsule encapsulating engineered bacteria. By sensing the confinement, the bacteria undergo programmed partial lysis at a high local density. Conversely, the encapsulating material shrinks responding to the changing chemical environment caused by cell growth, squeezing out the protein products released by bacterial lysis. This platform is then integrated with downstream modules to enable quantification of enzymatic kinetics, purification of diverse proteins, quantitative control of protein interactions and assembly of functional protein complexes and multienzyme metabolic pathways. Our work demonstrates the use of the cell–material feedback to engineer a modular and flexible platform with sophisticated yet well-defined programmed functions.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1: Programmable cell–material feedback for versatile protein production.
Fig. 2: Sustained synthesis and quantification of BlaM.
Fig. 3: Integrated protein production and purification.
Fig. 4: Composition control and DOL using MSBs.
Fig. 5: One-pot reconstruction of the multienzyme metabolic pathway underlying FAS.

Data availability

All data generated or analyzed during the current study are available from the corresponding author on reasonable request.

Code availability

All code used in this study is available from the author upon reasonable request.


  1. 1.

    Baeshen, M. N. et al. Production of biopharmaceuticals in E. coli: current scenario and future perspectives. J. Microbiol. Biotechnol. 25, 953–962 (2015).

    CAS  Article  Google Scholar 

  2. 2.

    Walsh, G. Biopharmaceutical benchmarks 2014. Nat. Biotechnol. 32, 992–1000 (2014).

    CAS  Article  Google Scholar 

  3. 3.

    Gottschalk, U., Brorson, K. & Shukla, A. A. The need for innovation in biomanufacturing. Nat. Biotechnol. 30, 489–492 (2012).

    CAS  Article  Google Scholar 

  4. 4.

    Dove, A. Uncorking the biomanufacturing bottleneck. Nat. Biotechnol. 20, 777–779 (2002).

    CAS  Article  Google Scholar 

  5. 5.

    Patient-centered drug manufacture. Nat. Biotechnol. 35, 485 (2017).

  6. 6.

    Schellekens, H., Aldosari, M., Talsma, H. & Mastrobattista, E. Making individualized drugs a reality. Nat. Biotechnol. 35, 507–513 (2017).

    CAS  Article  Google Scholar 

  7. 7.

    Choi, E. J. & Ling, G. S. Battlefield medicine: paradigm shift for pharmaceuticals manufacturing. PDA J. Pharm. Sci. Technol. 68, 312 (2014).

    Article  Google Scholar 

  8. 8.

    Ashok, A., Brison, M. & LeTallec, Y. Improving cold chain systems: challenges and solutions. Vaccine 35, 2217–2223 (2017).

    Article  Google Scholar 

  9. 9.

    Shukla, A. & Gottschalk, U. Single-use disposable technologies for biopharmaceutical manufacturing. Trends Biotechnol. 31, 147–154 (2013).

    CAS  Article  Google Scholar 

  10. 10.

    Lopes, A. Single-use in the biopharmaceutical industry: a review of current technology impact, challenges and limitations. Food Bioprod. Process. 93, 98–114 (2015).

    Article  Google Scholar 

  11. 11.

    Schilling, E., Kamholz, A. & Yager, P. Cell lysis and protein extraction in a microfluidic device with detection by a fluorogenic enzyme assay. Anal. Chem. 74, 1798–1804 (2002).

    CAS  Article  Google Scholar 

  12. 12.

    Jacquemart, R. et al. A single-use strategy to enable manufacturing of affordable biologics. Comput. Struct. Biotechnol. J. 14, 309–318 (2016).

    CAS  Article  Google Scholar 

  13. 13.

    Allison, N. & Richards, J. Current status and future trends for disposable technology in the biopharmaceutical industry. J. Chem. Technol. Biotechnol. 89, 1283–1287 (2014).

    CAS  Article  Google Scholar 

  14. 14.

    Mergulhão, F. J., Summers, D. K. & Monteiro, G. A. Recombinant protein secretion in Escherichia coli. Biotechnol. Adv. 23, 177–202 (2005).

    Article  Google Scholar 

  15. 15.

    Green, E. R. & Mecsas, J. Bacterial secretion systems: an overview. Microbiol. Spectr. (2016).

  16. 16.

    Mergulhao, F. J. M. & Monteiro, G. A. Secretion capacity limitations of the Sec pathway in Escherichia coli. J. Microbiol. Biotechnol. 14, 128–133 (2004).

    CAS  Google Scholar 

  17. 17.

    Xiyu, J., Jan, K. & Anthony, C. J. Controlled lysis of bacteria. US patent 7892811B2 (2011).

  18. 18.

    Din, M. O. et al. Synchronized cycles of bacterial lysis for in vivo delivery. Nature 536, 81–85 (2016).

    CAS  Article  Google Scholar 

  19. 19.

    Pardee, K. et al. Portable, on-demand biomolecular manufacturing. Cell 167, 248–259 (2016).

    CAS  Article  Google Scholar 

  20. 20.

    Karig, D. K., Bessling, S., Thielen, P., Zhang, S. & Wolfe, J. Preservation of protein expression systems at elevated temperatures for portable therapeutic production. J. R. Soc. Interface 14, 20161039 (2017).

    Article  Google Scholar 

  21. 21.

    Underwood, K. A., Swartz, J. R. & Puglisi, J. D. Quantitative polysome analysis identifies limitations in bacterial cell-free protein synthesis. Biotechnol. Bioeng. 91, 425–435 (2005).

    CAS  Article  Google Scholar 

  22. 22.

    Stech, M., Quast, R. B., Sachse, R., Schulze, C. & Kubick, D. A. W. S. A continuous-exchange cell-free protein synthesis system based on extracts from cultured insect cells. PLoS One 9, e96635 (2014).

    Article  Google Scholar 

  23. 23.

    Gutowska, A. et al. Squeezing hydrogels for controlled oral drug delivery. J. Control. Release 48, 141–148 (1997).

    CAS  Article  Google Scholar 

  24. 24.

    Eichenbaum, G. M., Kiser, P. F., Dobrynin, A. V., Simon, S. A. & Needham, D. Investigation of the swelling response and loading of ionic microgels with drugs and proteins: the dependence on cross-link density. Macromolecules 32, 4867–4878 (1999).

    CAS  Article  Google Scholar 

  25. 25.

    Huang, X. & Brazel, C. S. On the importance and mechanisms of burst release in matrix-controlled drug delivery systems. J. Control. Release 73, 121–136 (2001).

    CAS  Article  Google Scholar 

  26. 26.

    Marguet, P., Tanouchi, Y., Spitz, E., Smith, C. & You, L. Oscillations by minimal bacterial suicide circuits reveal hidden facets of host–circuit physiology. PLoS One 5, e11909 (2010).

    Article  Google Scholar 

  27. 27.

    Lopez-Leon, T., Carvalho, E., Seijo, B., Ortega-Vinuesa, J. & Bastos-Gonzalez, D. Physicochemical characterization of chitosan nanoparticles: electrokinetic and stability behavior. J. Colloid Interface Sci. 283, 344–351 (2005).

    CAS  Article  Google Scholar 

  28. 28.

    Gu, Z. et al. Glucose-responsive microgels integrated with enzyme nanocapsules for closed-loop insulin delivery. ACS Nano 7, 6758–6766 (2013).

    CAS  Article  Google Scholar 

  29. 29.

    Jonassen, H., Kjoniksen, A. L. & Hiorth, M. Effects of ionic strength on the size and compactness of chitosan nanoparticles. Colloid Polym. Sci. 290, 919–929 (2012).

    CAS  Article  Google Scholar 

  30. 30.

    Balagaddé, F. K., You, L., Hansen, C. L., Arnold, F. H. & Quake, S. R. Long-term monitoring of bacteria undergoing programmed population control in a microchemostat. Science 309, 137–140 (2005).

    Article  Google Scholar 

  31. 31.

    Tanouchi, Y., Pai, A., Buchler, N. E. & You, L. Programming stress-induced altruistic death in engineered bacteria. Mol. Syst. Biol. 8, 626 (2012).

    Article  Google Scholar 

  32. 32.

    Sun, F., Zhang, W. B., Mahdavi, A., Arnold, F. H. & Tirrell, D. A. Synthesis of bioactive protein hydrogels by genetically encoded SpyTag–SpyCatcher chemistry. Proc. Natl Acad. Sci. USA 111, 11269–11274 (2014).

    CAS  Article  Google Scholar 

  33. 33.

    Meyer, D. E. & Chilkoti, A. Purification of recombinant proteins by fusion with thermally-responsive polypeptides. Nat. Biotechnol. 17, 1112–1115 (1999).

    CAS  Article  Google Scholar 

  34. 34.

    Kwon, K. et al. High quality protein microarray using in situ protein purification. BMC Biotechnol. 9, 72 (2009).

    Article  Google Scholar 

  35. 35.

    Ge, X., Trabbic-Carlson, K., Chilkoti, A. & Filipe, C. D. Purification of an elastin-like fusion protein by microfiltration. Biotechnol. Bioeng. 95, 424–432 (2006).

    CAS  Article  Google Scholar 

  36. 36.

    Amiram, M., Luginbuhl, K. M., Li, X., Feinglos, M. N. & Chilkoti, A. Injectable protease-operated depots of glucagon-like peptide-1 provide extended and tunable glucose control. Proc. Natl Acad. Sci. USA 110, 2792–2797 (2013).

    CAS  Article  Google Scholar 

  37. 37.

    Lusvarghi, S. & Bewley, C. A. Griffithsin: an antiviral lectin with outstanding therapeutic potential. Viruses 8, 296 (2016).

    Article  Google Scholar 

  38. 38.

    Amato, G. et al. Recombinant growth hormone (GH) therapy in GH-deficient adults: a long-term controlled study on daily versus thrice weekly injections. J. Clin. Endocrinol. Metab. 85, 3720–3725 (2000).

    CAS  Article  Google Scholar 

  39. 39.

    Ihssen, J. et al. Production of glycoprotein vaccines in Escherichia coli. Microb. Cell Fact 9, 61 (2010).

    Article  Google Scholar 

  40. 40.

    Tsoi, R. et al. Metabolic division of labor in microbial systems. Proc. Natl Acad. Sci. USA 115, 2526–2531 (2018).

    CAS  Article  Google Scholar 

  41. 41.

    Villarreal, F. et al. Synthetic microbial consortia enable rapid assembly of pure translation machinery. Nat. Chem. Biol. 14, 29–35 (2018).

    CAS  Article  Google Scholar 

  42. 42.

    Scott, S. R. et al. A stabilized microbial ecosystem of self-limiting bacteria using synthetic quorum-regulated lysis. Nat. Microbiol. 2, 17083 (2017).

    CAS  Article  Google Scholar 

  43. 43.

    Xu, P. et al. Modular optimization of multi-gene pathways for fatty acids production in E. coli. Nat. Commun. 4, 1409 (2013).

    Article  Google Scholar 

  44. 44.

    Yu, X., Liu, T., Zhu, F. & Khosla, C. In vitro reconstitution and steady-state analysis of the fatty acid synthase from Escherichia coli. Proc. Natl Acad. Sci. USA 108, 18643–18648 (2011).

    CAS  Article  Google Scholar 

  45. 45.

    Heath, R. J. & Rock, C. O. Enoyl-acyl carrier protein reductase (fabI) plays a determinant role in completing cycles of fatty acid elongation in Escherichia coli. J. Biol. Chem. 270, 26538–26542 (1995).

    CAS  Article  Google Scholar 

  46. 46.

    Bi, H. K., Christensen, Q. H., Feng, Y. J., Wang, H. H. & Cronan, J. E. The Burkholderia cenocepacia BDSF quorum sensing fatty acid is synthesized by a bifunctional crotonase homologue having both dehydratase and thioesterase activities. Mol. Microbiol. 83, 840–855 (2012).

    CAS  Article  Google Scholar 

  47. 47.

    Pardee, K. et al. Paper-based synthetic gene networks. Cell 159, 940–954 (2014).

    CAS  Article  Google Scholar 

  48. 48.

    Lopatkin, A. J. & You, L. Synthetic biology looks good on paper. Cell 159, 718–720 (2014).

    CAS  Article  Google Scholar 

  49. 49.

    Perez-Pinera, P. et al. Synthetic biology and microbioreactor platforms for programmable production of biologics at the point-of-care. Nat. Commun. 7, 12211 (2016).

    CAS  Article  Google Scholar 

  50. 50.

    Hoare, T. R. & Kohane, D. S. Hydrogels in drug delivery: progress and challenges. Polymer 49, 1993–2007 (2008).

    CAS  Article  Google Scholar 

  51. 51.

    Worthington, A. S., Rivera, H., Torpey, J. W., Alexander, M. D. & Burkart, M. D. Mechanism-based protein cross-linking probes to investigate carrier protein-mediated biosynthesis. ACS Chem. Biol. 1, 687–691 (2006).

    CAS  Article  Google Scholar 

  52. 52.

    Lopatkin, A. J. et al. Antibiotics as a selective driver for conjugation dynamics. Nat. Microbiol. 1, 16044 (2016).

    CAS  Article  Google Scholar 

  53. 53.

    Huang, S. et al. Coupling spatial segregation with synthetic circuits to control bacterial survival. Mol. Syst. Biol. 12, 859 (2016).

    Article  Google Scholar 

  54. 54.

    Yu, T. et al. Reprogramming yeast metabolism from alcoholic fermentation to lipogenesis. Cell 174, 1549–1558 (2018).

    CAS  Article  Google Scholar 

Download references


We thank M. Lynch for plasmid constructs, insightful comments and suggestions; D.A. Tirrell, Y. Zhang and F. Sun for plasmid constructs; K. Zhu and K. Luginbuhl for insightful suggestions; J. Decker for useful suggestions; and P. Li, B. Chen, Q. Hu, X. Peng and Y. Zhang for assistance in revision. This study was partially supported by the U.S. Army Research Office under grant W911NF-14-1-0490 (to L.Y.), the National Institutes of Health (grant R01-GM098642 to L.Y. and grant R35GM127042 to A.C.), the Office of Naval Research (grant N00014-12-1-0631 to L.Y.), Beijing Municipal Natural Science Foundation (grant 5182017 to Z.L.) and a David and Lucile Packard Fellowship to L.Y.

Author information




Z.D. conceived the research, designed and performed experiments, interpreted the results, assisted in the model development and wrote the manuscript. A.J.L. developed the model and assisted in experimental setup, data interpretation and manuscript revisions. S.R. assisted in performing experiments, data analysis and manuscript revisions. T.A.S. and S.H. assisted in experimental setup, data interpretation and manuscript revisions. M.D. and X.Y. assisted in performing experiments, data interpretation and manuscript revisions. X.Z. and Z.L. assisted in performing experiments during the manuscript revisions. A.C. assisted in research design, experimental setup, data interpretation and manuscript revisions. L.Y. conceived the research and assisted in research design, modeling, data interpretation and manuscript writing.

Corresponding author

Correspondence to Lingchong You.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary information

Supplementary Tables 1–3 and Supplementary Figures 1–18

Reporting Summary

Supplementary Video 1

Oscillatory behavior of engineered cells was observed on the microfluidic device.

Supplementary Video 2

Capsules carrying engineered bacteria shrunk with cell growth.

Supplementary Video 3

Capsules carrying engineered bacteria swelled slightly before shrinking with cell growth.

Supplementary Video 4

Oscillations of the capsule size in the culture chamber were observed under microscopy.

Supplementary Video 5

Periodic accumulation of fluorescence in the assay chamber were observed under microscopy.

Supplementary Video 6

Response of chitosan capsules to periodic change in pH.

Supplementary Video 7

Alginate capsules maintain the same size at varied chemical conditions.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Dai, Z., Lee, A.J., Roberts, S. et al. Versatile biomanufacturing through stimulus-responsive cell–material feedback. Nat Chem Biol 15, 1017–1024 (2019).

Download citation

Further reading