阿维链霉菌中转录因子AveR对阿维菌素生物合成的正调节

Positive Regulation of Avermectin Biosynthesis by AveR in Streptomyces avermitilis

aveR是阿维链霉菌NRRL 8165阿维菌素生物合成基因簇中唯一可能的调节基因.为了验证aveR是否参与阿维菌素生物合成基因的转录调节,构建了用于敲除aveR的基因置换质粒pJTU2530,并通过接合转移引入了阿维链霉菌.通过筛选ThioSAprR转化子,经聚合酶链式反应(PCR)扩增验证,获得了aveR内部1 320 bp区域被阿泊拉霉素抗性基因aac(3)IV替换的突变株ZD10.高压液相色谱检测表明,与野生型菌株相比,突变株ZD10不再产生阿维菌素,并且ZD10中寡霉素的产量明显高于野生型菌株.进一步的反转录PCR(RT-PCR)分析表明,与野生型菌株相比,突变株ZD10的聚酮合酶基因aveA3不再转录.结果显示,AveR是阿维菌素生物合成的正调节因子,通过调节结构基因的转录表达来影响阿维菌素的产生.

Gene aver is the only putative regulatory gene in avermectin biosynthetic gene cluster from Streptomyces avermitilis NRRL8165. In order to test if aveR is involved in the regulation of transcription of avermectin biosynthetic genes, plasmid pJTU2530 was constructed and introduced into S. avermitilis by conjugation. The aver mutant ZD10, in which a 1320 bp internal fragment of aver is replaced by aac （3）IV, was obtained and confirmed through PCR amplification. Analyzed by LC-MS, avermectin productivity was abolished, whereas the yield of oligomycin significantly increased in ZD10. The reverse transcription PCR analysis indicates that the transcription level of aveA3, an essential PKS gene, is drastically reduced in ZD10. Therefore, AveR is proved as a positive regulator of avermeetin biosynthesis.