Kim Myoun-Su, Cho Wan-Je, Song Myoung Chong, Park Seong-Whan, Kim Kaeun, Kim Eunji, Lee Naryeong, Nam Sang-Jip, Oh Ki-Hoon, Yoon Yeo Joon
Department of Chemistry and Nano Science, Ewha Womans University, 52, Ewhayeodae-gil, Seoul, 03760, Republic of Korea.
Crop Protection R&D Center, FarmHannong Co., Ltd, 39-23, Dongan-ro 1113beon-gil, Yeonmu-eup, Nonsan-si, Chungcheongnam-do, 33010, Republic of Korea.
Microb Cell Fact. 2017 Jan 17;16(1):9. doi: 10.1186/s12934-017-0626-8.
Milbemycins, produced from Streptomyces hygroscopicus subsp. aureolacrimosus and Streptomyces bingchenggensis, are 16-membered macrolides that share structural similarity with avermectin produced from Streptomyces avermitilis. Milbemycins possess strong acaricidal, insecticidal, and anthelmintic activities but low toxicity. Due to the high commercial value of the milbemycins and increasing resistance to the avermectins and their derivatives, it is imperative to develop an efficient combinatorial biosynthesis system exploiting an overproduction host strain to produce the milbemycins and novel analogs in large quantities.
The respective replacement of AveA1 and AveA3 (or module 7 in AveA3) of the avermectin polyketide synthase (PKS) in the avermectin high-producing strain S. avermitilis SA-01 with MilA1 and MilA3 (or module 7 in MilA3) of the milbemycin PKS resulted in the production of milbemycins A3, A4, and D in small amounts and their respective C5-O-methylated congener milbemycins B2, B3, and G as major products with total titers of approximately 292 mg/l. Subsequent inactivation of the C5-O-methyltransferase AveD led to a production of milbemycins A3/A4 (the main components of the commercial product milbemectin) in approximately 225 and 377 mg/l in the flask and 5 l fermenter culture, respectively, along with trace amounts of milbemycin D.
We demonstrated that milbemycin biosynthesis can be engineered in the avermectin-producing S. avermitilis by combinatorial biosynthesis with only a slight decrease in its production level. Application of a similar strategy utilizing higher producing industrial strains will provide a more efficient combinatorial biosynthesis system based on S. avermitilis for further enhanced production of the milbemycins and their novel analogs with improved insecticidal potential.
由吸水链霉菌金色泪滴亚种和泾阳链霉菌产生的米尔倍霉素是16元大环内酯类化合物，与阿维链霉菌产生的阿维菌素结构相似。米尔倍霉素具有很强的杀螨、杀虫和驱虫活性，但毒性较低。由于米尔倍霉素具有很高的商业价值，且对阿维菌素及其衍生物的耐药性不断增加，因此必须开发一种高效的组合生物合成系统，利用过量生产宿主菌株大量生产米尔倍霉素和新型类似物。
用米尔倍霉素聚酮合酶的MilA1和MilA3（或MilA3中的模块7）分别替换阿维菌素高产菌株阿维链霉菌SA-01中阿维菌素聚酮合酶（PKS）的AveA1和AveA3（或AveA3中的模块7），导致少量米尔倍霉素A3、A4和D的产生，以及它们各自的C5-O-甲基化同系物米尔倍霉素B2、B3和G作为主要产物，总滴度约为292mg/L。随后C5-O-甲基转移酶AveD的失活导致在摇瓶和5L发酵罐培养中分别产生约225mg/L和377mg/L的米尔倍霉素A3/A4（商业产品米尔贝霉素的主要成分），以及微量的米尔倍霉素D。
我们证明了通过组合生物合成可以在产生阿维菌素的阿维链霉菌中对米尔倍霉素生物合成进行工程改造，其产量水平仅略有下降。利用更高产的工业菌株应用类似策略将提供一个基于阿维链霉菌的更高效组合生物合成系统，以进一步提高米尔倍霉素及其具有更高杀虫潜力的新型类似物的产量。
