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Discovery, Biosynthesis, and Rational Engineering of Novel Enterocin and Wailupemycin Polyketide Analogues

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Metabolomics Tools for Natural Product Discovery

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1055))

Abstract

The marine actinomycete Streptomyces maritimus produces a structurally diverse set of unusual polyketide natural products including the major metabolite enterocin. Investigations of enterocin biosynthesis revealed that the unique carbon skeleton is derived from an aromatic polyketide pathway which is genetically coded by the 21.3 kb enc gene cluster in S. maritimus. Characterization of the enc biosynthesis gene cluster and subsequent manipulation of it via heterologous expression and/or mutagenesis enabled the discovery of other enc-based metabolites that were produced in only very minor amounts in the wild type. Also described are techniques used to harness the enterocin biosynthetic machinery in order to generate unnatural enc-derived polyketide analogues. This review focuses upon the molecular methods used in combination with classical natural products detection and isolation techniques to access minor metabolites of the S. maritimus secondary metabolome.

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References

  1. Demain AL, Sanchez S (2009) Microbial drug discovery: 80 years of progress. J Antibiot 62:5–16

    Article  PubMed  CAS  Google Scholar 

  2. Omura S, Ikeda H, Ishikawa J, Hanamoto A, Takahashi C, Shinose M, Takahashi Y, Horikawa H, Nakazawa H, Osonoe T, Kikuchi H, Shiba T, Sakaki Y, Hattori M (2001) Genome sequence of an industrial microorganism Streptomyces avermitilis: deducing the ability of producing secondary metabolites. Proc Natl Acad Sci USA 98:12215–12220

    Article  PubMed  CAS  Google Scholar 

  3. Bentley SD, Chater KF, Cerdeno-Tarraga AM, Challis GL, Thomson NR, James KD, Harris DE, Quail MA, Kieser H, Harper D, Bateman A, Brown S, Chandra G, Chen CW, Collins M, Cronin A, Fraser A, Goble A, Hidalgo J, Hornsby T, Howarth S, Huang CH, Kieser T, Larke L, Murphy L, Oliver K, O’Neil S, Rabbinowitsch E, Rajandream MA, Rutherford K, Rutter S, Seeger K, Saunders D, Sharp S, Squares R, Squares S, Taylor K, Warren T, Wietzorrek A, Woodward J, Barrell BG, Parkhill J, Hopwood DA (2002) Complete genome sequence of the model actinomycete Streptomyces coelicolor A3(2). Nature 417:141–147

    Article  PubMed  Google Scholar 

  4. Ikeda H, Ishikawa J, Hanamoto A, Shinose M, Kikuchi H, Shiba T, Sakaki Y, Hattori M, Omura S (2003) Complete genome sequence and comparative analysis of the industrial microorganism Streptomyces avermitilis. Nat Biotechnol 21:526–531

    Article  PubMed  Google Scholar 

  5. Udwary DW, Zeigler L, Asolkar RN, Singan V, Lapidus A, Fenical W, Jensen PR, Moore BS (2007) Genome sequencing reveals complex secondary metabolome in the marine actinomycete Salinispora tropica. Proc Natl Acad Sci USA 104:10376–10381

    Article  PubMed  CAS  Google Scholar 

  6. Krug D, Zurek G, Revermann O, Vos M, Velicer GJ, Muller R (2008) Discovering the hidden secondary metabolome of Myxococcus xanthus: a study of intraspecific diversity. Appl Environ Microbiol 74:3058–3068

    Article  PubMed  CAS  Google Scholar 

  7. Winter JM, Behnken S, Hertweck C (2011) Genomics-inspired discovery of natural products. Curr Opin Chem Biol 15:22–31

    Article  PubMed  CAS  Google Scholar 

  8. Donadio S, Monciardini P, Sosio M (2007) Polyketide synthases and nonribosomal peptide synthetases: the emerging view from bacterial genomics. Nat Prod Rep 24:1073–1109

    Article  PubMed  CAS  Google Scholar 

  9. Fischbach MA, Walsh CT (2006) Assembly-line enzymology for polyketide and nonribosomal peptide antibiotics: logic, machinery, and mechanisms. Chem Rev 106:3468–3496

    Article  PubMed  CAS  Google Scholar 

  10. Nett M, Ikeda H, Moore BS (2009) Genomic basis for natural product biosynthetic diversity in the actinomycetes. Nat Prod Rep 26:1362–1384

    Article  PubMed  CAS  Google Scholar 

  11. Van Lanen SG, Shen B (2006) Microbial genomics for the improvement of natural product discovery. Curr Opin Microbiol 9:252–260

    Article  PubMed  Google Scholar 

  12. Lautru S, Deeth RJ, Bailey LM, Challis GL (2005) Discovery of a new peptide natural product by Streptomyces coelicolor genome mining. Nat Chem Biol 1:265–269

    Article  PubMed  CAS  Google Scholar 

  13. Kalaitzis JA, Lauro FM, Neilan BA (2009) Mining cyanobacterial genomes for genes encoding complex biosynthetic pathways. Nat Prod Rep 26:1447–1465

    Article  PubMed  CAS  Google Scholar 

  14. Walsh CT, Fischbach MA (2010) Natural products version 2.0: connecting genes to molecules. J Am Chem Soc 132:2469–2493

    Article  PubMed  CAS  Google Scholar 

  15. Piel J, Hoang K, Moore BS (2000) Natural metabolic diversity encoded by the enterocin biosynthesis gene cluster. J Am Chem Soc 122:5415–5416

    Article  CAS  Google Scholar 

  16. Piel J, Hertweck C, Shipley PR, Hunt DM, Newman MS, Moore BS (2000) Cloning, sequencing and analysis of the enterocin biosynthesis gene cluster from the marine isolate ‘Streptomyces maritimus’: evidence for the derailment of an aromatic polyketide synthase. Chem Biol 7:943–955

    Article  PubMed  CAS  Google Scholar 

  17. Staunton J, Weissman KJ (2001) Polyketide biosynthesis: a millennium review. Nat Prod Rep 18:380–416

    Article  PubMed  CAS  Google Scholar 

  18. Hertweck C (2009) The biosynthetic logic of polyketide diversity. Angew Chem Int Ed 48:4688–4716

    Article  CAS  Google Scholar 

  19. Austin MB, Noel JP (2003) The chalcone synthase superfamily of type III polyketide synthases. Nat Prod Rep 20:79–110

    Article  PubMed  CAS  Google Scholar 

  20. Khosla C, Kapur S, Cane DE (2009) Revisiting the modularity of modular polyketide synthases. Curr Opin Chem Biol 13:135–143

    Article  PubMed  CAS  Google Scholar 

  21. Shen B (2003) Polyketide biosynthesis beyond the type I, II and III polyketide synthase paradigms. Curr Opin Chem Biol 7:285–295

    Article  PubMed  CAS  Google Scholar 

  22. Van Lanen SG, Shen B (2008) Advances in polyketide synthase structure and function. Curr Opin Drug Disc 11:186–195

    Google Scholar 

  23. Hopwood DA (1997) Genetic contributions to understanding polyketide synthases. Chem Rev 97:2465–2497

    Article  PubMed  CAS  Google Scholar 

  24. Miyairi N, Sakai HI, Konomi T, Imanaka H (1976) Enterocin, a new antibiotic - taxonomy, isolation and characterization. J Antibiot 29:227–235

    Article  PubMed  CAS  Google Scholar 

  25. Tokuma Y, Miyairi N, Morimoto Y (1976) Structure of enterocin - x-ray analysis of m-bromobenzoyl enterocin dihydrate. J Antibiot 29:1114–1116

    Article  PubMed  CAS  Google Scholar 

  26. Seto H, Sato T, Urano S, Uzawa J, Yonehara H (1976) Utilization of C-13-C-13 coupling in structural and biosynthetic studies XII. Structure and biosynthesis of vulgamycin. Tetrahedron Lett 17:4367–4370

    Article  Google Scholar 

  27. Kang H, Jensen PR, Fenical W (1996) Isolation of microbial antibiotics from a marine ascidian of the genus Didemnum. J Org Chem 61:1543–1546

    Article  CAS  Google Scholar 

  28. Sitachitta N, Gadepalli M, Davidson BS (1996) New alpha-pyrone-containing metabolites from a marine-derived actinomycete. Tetrahedron 52:8073–8080

    Article  CAS  Google Scholar 

  29. Yu TW, Shen YM, McDaniel R, Floss HG, Khosla C, Hopwood DA, Moore BS (1998) Engineered biosynthesis of novel polyketides from Streptomyces spore pigment polyketide synthases. J Am Chem Soc 120:7749–7759

    Article  CAS  Google Scholar 

  30. Shen YM, Yoon P, Yu TW, Floss HG, Hopwood D, Moore BS (1999) Ectopic expression of the minimal whiE polyketide synthase generates a library of aromatic polyketides of diverse sizes and shapes. Proc Natl Acad Sci USA 96:3622–3627

    Article  PubMed  CAS  Google Scholar 

  31. Ziermann R, Betlach MC (1999) Recombinant polyketide synthesis in Streptomyces: engineering of improved host strains. Biotechniques 26:106–109

    PubMed  CAS  Google Scholar 

  32. Ziermann R, Betlach MC (2000) A two-vector system for the production of recombinant polyketides in Streptomyces. J Ind Microbiol Biot 24:46–50

    Article  CAS  Google Scholar 

  33. Alvarez MA, Fu H, Khosla C, Hopwood DA, Bailey JE (1996) Engineered biosynthesis of novel polyketides: properties of the whiE aromatase/cyclase. Nat Biotechnol 14:335–338

    Article  PubMed  CAS  Google Scholar 

  34. Xiang L, Kalaitzis JA, Nilsen G, Chen L, Moore BS (2002) Mutational analysis of the enterocin Favorskii biosynthetic rearrangement. Org Lett 4:957–960

    Article  PubMed  CAS  Google Scholar 

  35. Zhang HL, He XG, Adefarati A, Gallucci J, Cole SP, Beale JM, Keller PJ, Chang CJ, Floss HG (1990) Mutactin, a novel polyketide from Streptomyces coelicolor - structure and biosynthetic relationship to actinorhodin. J Org Chem 55:1682–1684

    Article  CAS  Google Scholar 

  36. McDaniel R, Ebert Khosla S, Fu H, Hopwood DA, Khosla C (1994) Engineered biosynthesis of novel polyketides - influence of a downstream enzyme on the catalytic specificity of a minimal aromatic polyketide synthase. Proc Natl Acad Sci USA 91:11542–11546

    Article  PubMed  CAS  Google Scholar 

  37. Graziani EI, Ritacco FV, Bernan VS, Telliez JB (2005) Phaeochromycins A-E, anti-inflammatory polyketides isolated from the soil actinomycete Streptomyces phaeochromogenes LL-P018. J Nat Prod 68:1262–1265

    Article  PubMed  CAS  Google Scholar 

  38. Hertweck C, Moore BS (2000) A plant-like biosynthesis of benzoyl-CoA in the marine bacterium ‘Streptomyces maritimus’. Tetrahedron 56:9115–9120

    Article  CAS  Google Scholar 

  39. Xiang L, Moore BS (2002) Inactivation, complementation, and heterologous expression of encP, a novel bacterial phenylalanine ammonia lyase gene. J Biol Chem 277:32505–32509

    Article  PubMed  CAS  Google Scholar 

  40. Xiang L, Moore BS (2003) Characterization of benzoyl coenzyme A biosynthesis genes in the enterocin-producing bacterium Streptomyces maritimus. J Bacteriol 185:399–404

    Article  PubMed  CAS  Google Scholar 

  41. Xiang L, Moore BS (2005) Biochemical characterization of a prokaryotic phenylalanine ammonia lyase. J Bacteriol 187:4286–4289

    Article  PubMed  CAS  Google Scholar 

  42. Izumikawa M, Cheng Q, Moore BS (2006) Priming type II polyketide synthases via a type II nonribosomal peptide synthetase mechanism. J Am Chem Soc 128:1428–1429

    Article  PubMed  CAS  Google Scholar 

  43. Kalaitzis JA, Izumikawa M, Xiang L, Hertweck C, Moore BS (2003) Mutasynthesis of enterocin and wailupemycin analogues. J Am Chem Soc 125:9290–9291

    Article  PubMed  CAS  Google Scholar 

  44. Hertweck C, Xiang L, Kalaitzis JA, Cheng Q, Palzer M, Moore BS (2004) Context-dependent behavior of the enterocin iterative polyketide synthase: a new model for ketoreduction. Chem Biol 11:461–468

    Article  PubMed  CAS  Google Scholar 

  45. Kalaitzis JA, Cheng Q, Meluzzi D, Xiang L, Izumikawa M, Dorrestein PC, Moore BS (2011) Policing starter unit selection of the enterocin type II polyketide synthase by the type II thioesterase EncL. Bioorgan Med Chem 19:6633–6638

    Article  CAS  Google Scholar 

  46. Xiang L, Kalaitzis JA, Moore BS (2004) EncM, a versatile enterocin biosynthetic enzyme involved in Favorskii oxidative rearrangement, aldol condensation, and heterocycle-forming reactions. Proc Natl Acad Sci USA 101:15609–15614

    Article  PubMed  CAS  Google Scholar 

  47. Werneburg M, Busch B, He J, Richter MEA, Xiang L, Moore BS, Roth M, Dahse HM, Hertweck C (2010) Exploiting enzymatic promiscuity to engineer a focused library of highly selective antifungal and antiproliferative aureothin analogues. J Am Chem Soc 132:10407–10413

    Article  PubMed  CAS  Google Scholar 

  48. Cheng Q, Xiang L, Izumikawa M, Meluzzi D, Moore BS (2007) Enzymatic total synthesis of enterocin polyketides. Nat Chem Biol 3:557–558

    Article  PubMed  CAS  Google Scholar 

  49. Kalaitzis JA, Cheng Q, Thomas PM, Kelleher NL, Moore BS (2009) In vitro biosynthesis of unnatural enterocin and wailupemycin polyketides. J Nat Prod 72:469–472

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

I wish to thank Professor Bradley S. Moore (Scripps Institution of Oceanography, UCSD) for providing me with the opportunity to work on the enterocin project, and also for his advice, support, and mentorship. I also thank many colleagues and collaborators who are referenced throughout this paper for their vast contributions.

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Authors and Affiliations

  1. School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, Australia

    John A. Kalaitzis

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  1. John A. Kalaitzis
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Editors and Affiliations

  1. Parkville, Australia

    Ute Roessner

  2. School of Botany, The University of Melbourne, Parkville, Victoria, Australia

    Daniel Anthony Dias

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Kalaitzis, J.A. (2013). Discovery, Biosynthesis, and Rational Engineering of Novel Enterocin and Wailupemycin Polyketide Analogues. In: Roessner, U., Dias, D. (eds) Metabolomics Tools for Natural Product Discovery. Methods in Molecular Biology, vol 1055. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-577-4_13

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  • DOI: https://doi.org/10.1007/978-1-62703-577-4_13

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-576-7

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