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Cell-free synthesis of polyketides by recombinant erythromycin polyketide synthases

Abstract

MODULAR polyketide synthases (PKSs) are complex multi-enzyme proteins that catalyse the bacterial biosynthesis of many pharma-ceutically useful polyketides. The PKSs are organized into a series of modules, each containing the active catalytic sites required for one step in the synthesis process1–4. Here we report a method for cell-free enzymatic synthesis of 6-deoxyerythronolide B (6-dEB), the parent molecule of the antibiotic erythromycin A, using recombinant 6-deoxyerythronolide B synthase (DEBS), a modular PKS with at least 28 distinct active sites. We have also synthesized in vitro a triketide lactone by using a truncated mutant of DEBS. The availability of such cell-free synthetic routes will allow direct investigation of the structural and mechanistic basis for the unusual combination of high substrate specificity5–10 and tolerance to genetic reprogramming2,11–15 found in this enzyme family.

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References

  1. Cortes, J., Haydock, S. F., Roberts, G. A., Bevitt, D. J. & Leadlay, P. F. Nature 348, 176–178 (1990).

    Article  ADS  CAS  Google Scholar 

  2. Donadio, S., Staver, M. J., McAlpine, J. B., Swanson, S. J. & Katz, L. Science 252, 675–679 (1991).

    Article  ADS  CAS  Google Scholar 

  3. MacNeil, D. J. et al. Gene 115, 119–125 (1992).

    Article  CAS  Google Scholar 

  4. Schwecke, T. et al. Proc. natn. Acad. Sci. U.S.A. 92, 7839–7843 (1995).

    Article  ADS  CAS  Google Scholar 

  5. Cane, D. E. & Yang, C. C. J. Am. chem. Soc. 109, 1255–1257 (1987).

    Article  CAS  Google Scholar 

  6. Yue, S., Duncan, J. S., Yamamoto, Y. & Hutchinson, C. R. J. Am. chem. Soc. 109, 1253–1255 (1987).

    Article  CAS  Google Scholar 

  7. Cane, D. E., Prabhakaran, P. C., Tan, W. & Ott, W. R. Tetrahedron Lett. 32, 5457–5460 (1991).

    Article  CAS  Google Scholar 

  8. Cane, D. E., Lambalot, R. H., Prabhakaran, P. C. & Ott, W. R. J. Am. chem. Soc. 115, 522–566 (1993).

    Article  CAS  Google Scholar 

  9. Dutton, C. J., Hooper, A. M., Leadlay, P. F. & Staunton, J. Tetrahedron Lett. 35, 327–330 (1994).

    Article  CAS  Google Scholar 

  10. Cane, D. E. & Luo, G. J. Am. chem. Soc. 117, 6633–6634 (1995).

    Article  CAS  Google Scholar 

  11. Donadio, S., McAlpine, J. B., Sheldon, P. J., Jackson, M. & Katz, L. Proc. natn. Acad. Sci. U.S.A. 90, 7119–7123 (1993).

    Article  ADS  CAS  Google Scholar 

  12. Kao, C. M., Luo, G., Katz, L., Cane, D. E. & Khosla, C. J. Am. chem. Soc. 116, 11612–11613 (1994).

    Article  CAS  Google Scholar 

  13. Kao, C. M., Luo, G., Katz, L., Cane, D. E. & Khosla, C. J. Am. chem. Soc. 117, 9105–9106 (1995).

    Article  Google Scholar 

  14. Cortes, J. et al. Science 268, 1487–1489 (1995).

    Article  ADS  CAS  Google Scholar 

  15. Brown, M. J. B., Cortes, J., Cutter, A. L., Leadlay, P. F. & Staunton, J. J. chem. Soc. chem. Commun. 1517–1518 (1995).

  16. Wakil, S. J. Biochemistry 28, 4523–4530 (1989).

    Article  CAS  Google Scholar 

  17. O'Hagan, D. The Polyketide Metabolites (Norwood, Chichester, 1991).

    Google Scholar 

  18. Katz, L. & Donadio, S. A. Rev. Microbiol. 47, 875–912 (1993).

    Article  CAS  Google Scholar 

  19. Kao, C. M., Katz, L. & Khosla, C. Science 265, 509–512 (1994).

    Article  ADS  CAS  Google Scholar 

  20. Marsden, A. F. A. et al. Science 263, 378–380 (1994).

    Article  ADS  CAS  Google Scholar 

  21. Wawszkiewicz, E. J. & Lynen, F. Biochem. Zbl. 340, 213–227 (1964).

    CAS  Google Scholar 

  22. Corcoran, J. W. in Antibiotics IV. Biosynthesis (ed. Corcoran, J. W.) 146–148 (Springer, New York, 1981).

    Google Scholar 

  23. Roberts, G. & Leadlay, P. F. Biochem. Soc. Trans. 12, 642–643 (1984).

    Article  CAS  Google Scholar 

  24. Hunaiti, A. A. & Kolattukudy, P. E. Antimicrob. Agents Chemother. 25, 173–178 (1984).

    Article  CAS  Google Scholar 

  25. Caffrey, P., Bevitt, D. J., Staunton, J. & Leadlay, P. F. FEBS Lett. 304, 225–228 (1992).

    Article  CAS  Google Scholar 

  26. Leadlay, P. F. et al. Biochem. Soc. Trans. 21, 218–222 (1993).

    Article  CAS  Google Scholar 

  27. Dimroth, P., Walter, H. & Lynen, F. Eur. J. Biochem. 13, 98–110 (1970).

    Article  CAS  Google Scholar 

  28. Lanz, T., Tropf, S., Marner, F. J., Schröder, J. & Schröder, G. J. biol. Chem. 266, 9971–9976 (1991).

    CAS  PubMed  Google Scholar 

  29. Shen, B. & Hutchinson, C. R. Science 262, 1535–1540 (1993).

    Article  ADS  CAS  Google Scholar 

  30. Offenzeller, M. et al. J. biol. Chem. 268, 26127–26134 (1993).

    CAS  PubMed  Google Scholar 

  31. Cane, D. E., Luo, G., Khosla, C., Kao, C. M. & Katz, L. J. Antibiot. 48, 647–651 (1995).

    Article  CAS  Google Scholar 

  32. Plate, C. A., Joshi, V. C. & Wakil, S. J. J. biol. Chem. 245, 2868–2875 (1970).

    CAS  PubMed  Google Scholar 

  33. D'Agnolo, G. et al. Biochim. biophys. Acta 326, 155–166 (1973).

    Article  CAS  Google Scholar 

  34. Pieper, R., Luo, G., Cane, D. E. & Khosla, C. J. Am. chem. Soc. (in the press).

  35. Wiesmann, K. E. H. et al. Chem. Biol. 2, 583–589 (1995).

    Article  CAS  Google Scholar 

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Pieper, R., Luo, G., Cane, D. et al. Cell-free synthesis of polyketides by recombinant erythromycin polyketide synthases. Nature 378, 263–266 (1995). https://doi.org/10.1038/378263a0

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