Heterologous production of kasugamycin, an aminoglycoside antibiotic from Streptomyces kasugaensis, in Streptomyces lividans and Rhodococcus erythropolis L-88 by constitutive expression of the biosynthetic gene cluster
Heterologous production of kasugamycin, an aminoglycoside antibiotic from Streptomyces kasugaensis, in Streptomyces lividans and Rhodococcus erythropolis L-88 by constitutive expression of the biosynthetic gene cluster
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DOI：
10.1007/s00253-017-8189-5
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发表时间：
2017-05-01
影响因子：
5
通讯作者：
Kojima, Ikuo
中科院分区：
文献类型：
--
作者：
Kasuga, Kano;Sasaki, Akira;Kojima, Ikuo
春雷霉素（KSM）是一种分离自春日链霉菌（Streptomyces kasugaensis）的氨基糖苷类抗生素，用于防治稻瘟病已有50多年的历史。我们克隆了来自S. kasugaensis MB 273-C4并构建了三个KBG盒（即，盒I-III），以使得能够通过KBG的组成型表达在许多放线菌中异源产生KSM。盒I包含簇中所有推定的转录单位，但它被置于来自Tn 5的P（neo）启动子的控制下。它不能在变铅青链霉菌中稳定存在，也不能转化红串红球菌。盒II保留了KBG的原始排列，除了KBG的特异性激活基因kasT的启动子被来自阿维链霉菌的rpsJ的组成型启动子P（rpsJ）取代。为了提高细胞内肌醇的浓度，我们构建了编码肌醇-1-磷酸合成酶的ino 1基因的表达盒。将阿维菌素插入盒II中以产生盒III。这两个盒在S. lividans和R.以生产KSM。特别地，S.变铅青聚糖诱导KSM的产生达到与S.春日此外，盒III比盒II在R中诱导更多的KSM积累。erythropolis，表明通过ino 1在宿主中的表达外源性供应肌醇。CascaseII和III似乎对放线菌中的异源KSM生产是有用的。红球菌在液体培养中表现出球形，也是一种很有前途的抗生素发酵异源宿主。
Kasugamycin (KSM), an aminoglycoside antibiotic isolated from Streptomyces kasugaensis cultures, has been used against rice blast disease for more than 50 years. We cloned the KSM biosynthetic gene (KBG) cluster from S. kasugaensis MB273-C4 and constructed three KBG cassettes (i.e., cassettes I-III) to enable heterologous production of KSM in many actinomycetes by constitutive expression of KBGs. Cassette I comprised all putative transcriptional units in the cluster, but it was placed under the control of the P (neo) promoter from Tn5. It was not maintained stably in Streptomyces lividans and did not transform Rhodococcus erythropolis. Cassette II retained the original arrangement of KBGs, except that the promoter of kasT, the specific activator gene for KBG, was replaced with P (rpsJ) , the constitutive promoter of rpsJ from Streptomyces avermitilis. To enhance the intracellular concentration of myo-inositol, an expression cassette of ino1 encoding the inositol-1-phosphate synthase from S. avermitilis was inserted into cassette II to generate cassette III. These two cassettes showed stable maintenance in S. lividans and R. erythropolis to produce KSM. Particularly, the transformants of S. lividans induced KSM production up to the same levels as those produced by S. kasugaensis. Furthermore, cassette III induced more KSM accumulation than cassette II in R. erythropolis, suggesting an exogenous supply of myo-inositol by the ino1 expression in the host. Cassettes II and III appear to be useful for heterologous KSM production in actinomycetes. Rhodococcus exhibiting a spherical form in liquid cultivation is also a promising heterologous host for antibiotic fermentation.