Increased heterologous production of the antitumoral polyketide mithramycin A by engineered Streptomyces lividans TK24 strains

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Abstract

Mithramycin A is an antitumor compound used for treatment of several types of cancer including chronic and acute myeloid leukemia, testicular carcinoma, hypercalcemia and Paget’s disease. Selective modifications of this molecule by combinatorial biosynthesis and biocatalysis opened the possibility to produce mithramycin analogues with improved properties that are currently under preclinical development. The mithramycin A biosynthetic gene cluster from Streptomyces argillaceus ATCC12956 was cloned by transformation assisted recombination in Saccharomyces cerevisiae and heterologous expression in Streptomyces lividans TK24 was evaluated. Mithramycin A was efficiently produced by S. lividans TK24 under standard fermentation conditions. To improve the yield of heterologously produced mithramycin A, a collection of derivative strains of S. lividans TK24 were constructed by sequential deletion of known potentially interfering secondary metabolite gene clusters using a protocol based on the positive selection of double crossover events with blue pigment indigoidine-producing gene. Mithramycin A production was evaluated in these S. lividans strains and substantially improved mithramycin A production was observed depending on the deleted gene clusters. A collection of S. lividans strains suitable for heterologous expression of actinomycetes secondary metabolites were generated and efficient production of mithramycin A with yields close to 3 g/L, under the tested fermentation conditions was achieved using these optimized collection of strains.

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Change history

  • 27 December 2017

    The original publication contains error error in the Materials and Methods section and in the acknowledgement section.

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Acknowledgements

This work was supported by the Slovak Research and Development Agency under contract APVV-15-0410 and by the VEGA grant 2/0002/16 from Slovak Academy of Sciences. The research leading to these results has received funding from the European Commission’s Seventh Framework Programme (FP7/2007-2013) under the grant agreement STREPSYNTH (project No. 613877). This work was co–funded by the Slovak Research and Development Agency under the contract No. DO7RP-0037-12. We are grateful to A. Luzhetskyy for providing cosmid 1443::AmR and S. Zotchev for providing plasmid pCLY10 and S. cerevisiae BY4742.

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Correspondence to Jesús Cortés or Jan Kormanec.

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A correction to this article is available online at https://doi.org/10.1007/s00253-017-8710-x.

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Novakova, R., Núñez, L.E., Homerova, D. et al. Increased heterologous production of the antitumoral polyketide mithramycin A by engineered Streptomyces lividans TK24 strains. Appl Microbiol Biotechnol 102, 857–869 (2018). https://doi.org/10.1007/s00253-017-8642-5

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Keywords

  • Mithramycin
  • Streptomyces lividans
  • TAR cloning
  • Heterologous expression