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Genetic engineering of Streptomyces bingchenggensis to produce milbemycins A3/A4 as main components and eliminate the biosynthesis of nanchangmycin

  • Applied genetics and molecular biotechnology
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Abstract

Milbemycins A3/A4 are important 16-membered macrolides which have been commercialized and widely used as pesticide and veterinary medicine. However, similar to other milbemycin producers, the production of milbemycins A3/A4 in Streptomyces bingchenggensis is usually accompanied with undesired by-products such as C5-O-methylmilbemycins B2/B3 (α-class) and β1/β2 (β-class) together with nanchangmycin. In order to obtain high yield milbemycins A3/A4-producing strains that produce milbemycins A3/A4 as main components, milD, a putative C5-O-methyltransferase gene of S. bingchenggensis, was biofunctionally investigated by heterologous expression in Escherichia coli. Enzymatic analysis indicated that MilD can catalyze both α-class (A3/A4) and β-class milbemycins (β11) into C5-O-methylmilbemycins B2/B3 and β1, respectively, suggesting little effect of furan ring formed between C6 and C8a on the C5-O-methylation catalyzed by MilD. Deletion of milD gene resulted in the elimination of C5-O-methylmilbemycins B2/B3 and β1/β2 together with an increased yield of milbemycins A3/A4 in disruption strain BCJ13. Further disruption of the gene nanLD encoding loading module of polyketide synthase responsible for the biosynthesis of nanchangmycin led to strain BCJ36 that abolished the production of nanchangmycin. Importantly, mutant strain BCJ36 (∆milD∆nanLD) produced milbemycins A3/A4 as main secondary metabolites with a yield of 2312 ± 47 μg/ml, which was approximately 74 % higher than that of the initial strain S. bingchenggensis BC-109-6 (1326 ± 37 μg/ml).

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Acknowledgments

This research was supported by the National Key Technology R&D Program (2010CB126102), the Outstanding Youth Foundation of China (31225024), the National Natural Science Foundation of China (30971937), the Program for New Century Excellent Talents in University (NCET-08-0668, 1154-NCET-002), the Outstanding Youth Foundation of Heilongjiang Province (JC200706), and the Program for New Teachers in University (20092325120007).

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Authors and Affiliations

  1. School of Life Science, Northeast Agricultural University, Harbin, 150030, People’s Republic of China

    Ji Zhang, Hai-Rong He & Wen-Sheng Xiang

  2. Key Laboratory of Microbial Drug Engineering of Heilongjiang Provincial Education Committee, Harbin, 150030, People’s Republic of China

    Jing An, Ji-Jia Wang, Yi-Jun Yan & Xiang-Jing Wang

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  1. Ji Zhang
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  2. Jing An
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Correspondence to Xiang-Jing Wang or Wen-Sheng Xiang.

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Zhang and An contributed equally to this work.

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Zhang, J., An, J., Wang, JJ. et al. Genetic engineering of Streptomyces bingchenggensis to produce milbemycins A3/A4 as main components and eliminate the biosynthesis of nanchangmycin. Appl Microbiol Biotechnol 97, 10091–10101 (2013). https://doi.org/10.1007/s00253-013-5255-5

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