Recombinatorial biosynthesis of polyketides

  • Antonio Starcevic
  • Kerstin Wolf
  • Janko Diminic
  • Jurica Zucko
  • Ida Trninic Ruzic
  • Paul F. Long
  • Daslav Hranueli
  • John Cullum
Genetics and Molecular Biology of Industrial Organisms


Modular polyketide synthases (PKSs) from Streptomyces and related genera of bacteria produce many important pharmaceuticals. A program called CompGen was developed to carry out in silico homologous recombination between gene clusters encoding PKSs and determine whether recombinants have cluster architectures compatible with the production of polyketides. The chemical structure of recombinant polyketides was also predicted. In silico recombination was carried out for 47 well-characterised clusters. The predicted recombinants would produce 11,796 different polyketide structures. The molecular weights and average degree of reduction of the chemical structures are dispersed around the parental structures indicating that they are likely to include pharmaceutically interesting compounds. The details of the recombinants and the chemical structures were entered in a database called r-CSDB. The virtual compound library is a useful resource for computer-aided drug design and chemoinformatics strategies for finding pharmaceutically relevant chemical entities. A strategy to construct recombinant Streptomyces strains to produce these polyketides is described and the critical steps of mobilizing large biosynthetic clusters and producing new linear cloning vectors are illustrated by experimental data.


Polyketides Actinobacteria Gene clusters Homologous recombination 



We thank David Hopwood for plasmid pIJ903 and Stanley N. Cohen for plasmid pQC48. This work was supported by a grant (058-0000000-3475 to D.H.) from the Ministry of Science, Education and Sports, Republic of Croatia and by a cooperation grant of the German Academic Exchange Service (DAAD) and the Ministry of Science, Education and Sports, Republic of Croatia (to D.H. and J.C.).

Supplementary material

10295_2011_1049_MOESM1_ESM.doc (341 kb)
Supplementary material 1 (DOC 341 kb)


  1. 1.
    Baltz RH (2006) Molecular engineering approaches to peptide, polyketide and other antibiotics. Nat Biotechnol 24:1533–1540PubMedCrossRefGoogle Scholar
  2. 2.
    Bieganski P, Riedl J, Carlis JV, Retzel EF (1994) Generalized suffix trees for biological sequence data. Proceedings of the twenty-seventh Hawaii international conference on system sciences, vol V (biotechnology computing), pp 35–44. doi:10.1109/HICSS.1994.323593
  3. 3.
    Bystrykh LV, Fernández-Moreno MA, Herrema JK, Malpartida F, Hopwood DA, Dijkhuizen L (1996) Production of actinorhodin-related “blue pigments” by Streptomyces coelicolor A3(2). J Bacteriol 178:2238–2244PubMedGoogle Scholar
  4. 4.
    Chen CW, Huang CH, Lee HH, Tsai HH, Kirby R (2002) Once the circle has been broken: dynamics and evolution of Streptomyces chromosomes. Trends Genet 18:522–529PubMedCrossRefGoogle Scholar
  5. 5.
    Cruz MC, Cavallo LM, Görlach JM, Cox G, Perfect JR, Cardenas ME, Heitman J (1999) Rapamycin antifungal action is mediated via conserved complexes with FKBP12 and TOR kinase homologs in Cryptococcus neoformans. Mol Cell Biol 19:4101–4112PubMedGoogle Scholar
  6. 6.
    Gibson DG, Glass JI, Lartigue C, Noskov VN, Chuang RY, Algire MA, Benders GA, Montague MG, Ma L, Moodie MM, Merryman C, Vashee S, Krishnakumar R, Assad-Garcia N, Andrews-Pfannkoch C, Denisova EA, Young L, Qi ZQ, Segall-Shapiro TH, Calvey CH, Parmar PP, Hutchison CA 3rd, Smith HO, Venter JC (2010) Creation of a bacterial cell controlled by a chemically synthesized genome. Science 329:52–56PubMedCrossRefGoogle Scholar
  7. 7.
    Hopwood DA, Kieser T, Wright HM, Bibb MJ (1983) Plasmids, recombination, and chromosomal mapping in Streptomyces lividans 66. J Gen Microbiol 129:2257–2269PubMedGoogle Scholar
  8. 8.
    Hosted TJ, Baltz RH (1997) Use of rpsL for dominance selection and gene replacement in Streptomyces roseosporus. J Bacteriol 179:180–186PubMedGoogle Scholar
  9. 9.
    Hranueli D, Cullum J, Basrak B, Goldstein P, Long PF (2005) Plasticity of the Streptomyces genome: evolution and engineering of new antibiotics. Curr Med Chem 12:1697–1704PubMedCrossRefGoogle Scholar
  10. 10.
    Jenke-Kodama H, Dittmann E (2009) Bioinformatic perspectives on NRPS/PKS megasynthases: advances and challenges. Nat Prod Rep 26:874–883PubMedCrossRefGoogle Scholar
  11. 11.
    Khosla C, Kapur S, Cane DE (2009) Revisiting the modularity of modular polyketide synthases. Curr Opin Chem Biol 13:135–143PubMedCrossRefGoogle Scholar
  12. 12.
    Kieser T, Bibb MJ, Buttner MJ, Hopwood DA (2000) Practical Streptomyces genetics. The John Innes Foundation, NorwichGoogle Scholar
  13. 13.
    Kinashi H, Shimaji-Murayama M, Hanafusa T (1992) Integration of SCP1, a giant linear plasmid, into the Streptomyces coelicolor chromosome. Gene 115:35–41PubMedCrossRefGoogle Scholar
  14. 14.
    Lydiate DJ, Malpartida F, Hopwood DA (1985) The Streptomyces plasmid SCP2*: its functional analysis and development into useful cloning vectors. Gene 35:223–235PubMedCrossRefGoogle Scholar
  15. 15.
    Malpartida F, Hopwood DA (1986) Physical and genetic characterisation of the gene cluster for the antibiotic actinorhodin in Streptomyces coelicolor A3(2). Mol Gen Genet 205:66–73PubMedCrossRefGoogle Scholar
  16. 16.
    Medema MH, Breitling R, Bovenberg R, Takano E (2011) Exploiting plug-and-play synthetic biology for drug discovery and production in microorganisms. Nat Rev Microbiol 9:131–137PubMedCrossRefGoogle Scholar
  17. 17.
    Needleman SB, Wunsch CD (1970) A general method applicable to the search for similarities in the amino acid sequence of two proteins. J Mol Biol 48:443–453PubMedCrossRefGoogle Scholar
  18. 18.
    Norrander J, Kempe T, Messing J (1983) Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene 26:101–106PubMedCrossRefGoogle Scholar
  19. 19.
    Pandza S, Biuković G, Paravic A, Dadbin A, Cullum J, Hranueli D (1998) Recombination between the linear plasmid pPZG101 and the linear chromosome of Streptomyces rimosus can lead to exchange of ends. Mol Microbiol 28:1165–1176PubMedCrossRefGoogle Scholar
  20. 20.
    Paravic A (2001) PhD dissertation, University of Kaiserslautern, KaiserslauternGoogle Scholar
  21. 21.
    Qin Z, Shen M, Cohen SN (2003) Identification and characterization of a pSLA2 plasmid locus required for linear DNA replication and circular plasmid stable inheritance in Streptomyces lividans. J Bacteriol 185:6575–6582PubMedCrossRefGoogle Scholar
  22. 22.
    Rice P, Longden I, Bleasby A (2000) EMBOSS: the European molecular biology open software suite. Trends Genet 2000(16):276–277CrossRefGoogle Scholar
  23. 23.
    Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory, Cold Spring HarborGoogle Scholar
  24. 24.
    Shen P, Huang HV (1986) Homologous recombination in Escherichia coli: dependence on substrate length and homology. Genetics 112:441–457PubMedGoogle Scholar
  25. 25.
    Starcevic A, Zucko J, Simunkovic J, Long PF, Cullum J, Hranueli D (2008) ClustScan: an integrated program package for the semi-automatic annotation of modular biosynthetic gene clusters and in silico prediction of novel chemical structures. Nucleic Acids Res 36:6882–6892PubMedCrossRefGoogle Scholar
  26. 26.
    Starcevic A, Diminic J, Zucko J, Elbekali M, Schlosser T, Lisfi M, Vukelic A, Long PF, Hranueli D, Cullum J (2011) A novel docking domain interface model that can predict recombination between homoeologous modular biosynthetic gene clusters. J Ind Microbiol Biotechnol. doi:10.1007/s10295-010-0909-0
  27. 27.
    Stoll A, Horvat LI, Lopes-Shikida SA, Padilla G, Cullum J (2000) Isolation and cloning of Streptomyces terminal fragments. Antonie Van Leeuwenhoek 78:223–226PubMedCrossRefGoogle Scholar
  28. 28.
    Weininger D (1988) SMILES, a chemical language and information system. 1. Introduction to methodology and encoding rules. J Chem Inf Comput Sci 28:31–36CrossRefGoogle Scholar
  29. 29.
    Xu M, Zhu Y, Zhang R, Shen M, Jiang W, Zhao G, Qin Z (2006) Characterization of the genetic components of Streptomyces lividans linear plasmid SLP2 for replication in circular and linear modes. J Bacteriol 188:6851–6857PubMedCrossRefGoogle Scholar
  30. 30.
    Zucko J, Cullum J, Hranueli D, Long PF (2011) Evolutionary dynamics of modular polyketide synthases, with implications for protein design and engineering. J Antibiot 64:89–92PubMedCrossRefGoogle Scholar

Copyright information

© Society for Industrial Microbiology 2011

Authors and Affiliations

  • Antonio Starcevic
    • 1
  • Kerstin Wolf
    • 2
  • Janko Diminic
    • 1
  • Jurica Zucko
    • 1
  • Ida Trninic Ruzic
    • 1
    • 4
  • Paul F. Long
    • 3
  • Daslav Hranueli
    • 1
  • John Cullum
    • 2
  1. 1.Faculty of Food Technology and BiotechnologyUniversity of ZagrebZagrebCroatia
  2. 2.Department of GeneticsUniversity of KaiserslauternKaiserslauternGermany
  3. 3.Institute of Pharmaceutical ScienceKing’s College LondonLondonUK
  4. 4.Novalis d.o.o.ZagrebCroatia

Personalised recommendations