卡伍尔氏链霉菌NA4生物合成巴弗洛霉素前体甲氧基丙二酰ACP来源的研究  

The Source of Methoxymalonyl-ACP Precursor for Bafilomycin Biosynthesis by Streptomyces cavourensis NA4

作　　者：鲍秀艳 姚雪春 史美君 胡江春[1] 潘华奇[1] BAO Xiu-yan;YAO Xue-chun;SHI Mei-jun;HU Jiang-chun;PAN Hua-qi(Shenyang Inst.of Appl.Ecol.,Chinese Acad.of Sci.,Shenyang 110016;Uni.of Chinese Acad.of Sci.,Beijing 100049)

机构地区：[1]中国科学院沈阳应用生态研究所,辽宁沈阳110016 [2]中国科学院大学,北京100049

出　　处：《微生物学杂志》2022年第1期9-16,共8页Journal of Microbiology

基　　金：国家自然科学基金项目(31872036);辽宁省“兴辽英才计划”项目(XLYC1807268);中国科学院青年创新促进会项目(2018229)。

摘　　要：巴弗洛霉素(bafilomycin)是一类由Ⅰ型聚酮合成酶装配合成的具有十六元大环内酯骨架的化合物,是研发农药和医药的良好母体。深海来源的卡伍尔氏链霉菌(Streptomyces cavourensis) NA4能产生巴弗洛霉素,而较低的发酵产量限制了巴弗洛霉素产业化应用。明确前体来源是理性构建高产菌株的基础,为了确定卡伍尔氏链霉菌NA4中合成巴弗洛霉素重要前体甲氧基丙二酰ACP的来源,构建了甲氧基丙二酰ACP生物合成部分基因bafB-E缺失突变株。实时荧光定量PCR分析发现野生株NA4中bafB mRNA表达水平随培养时间延长而不断增强,突变株则检测不到bafB mRNA的表达,显示ΔbafB-E突变株构建成功。进一步HPLC-UV分析结果显示,相比野生株NA4,ΔbafB-E突变株不再产生巴弗洛霉素。提示野生株NA4中bafB-F编码的合成途径是巴弗洛霉素装配前体甲氧基丙二酰ACP的唯一来源。将为通过强化前体供应策略理性构建巴弗洛霉素高产菌株提供参考。Bafilomycins are kinds of compound possessing 16-element macrolides skeleton, being assembled and synthesized by type Ⅰ polyketide synthase. It is a fine matrix for R&D of pesticides and medicine. And a deep-sea-derived actinomycete Streptomyces cavourensis NA4 could produce bafilomycin. However, low fermentation yield has restricted the industrial application of bafilomycin. Therefore, being explicit the source of the precursors is the basis of rational construction of high-yielding strains. In order to ascertain the source of the important precursor methoxymalonoyl ACP in the synthesis of bafilomycin, in this study a bafB-E gene cluster, a partial biosynthetic gene for methoxymalonoyl ACP, deleted mutant was constructed. Real-time PCR analysis showed that the mRNA expression of bafB in wild strain NA4 increased continuously with prolonging of culture time, while the mRNA expression of bafB was almost not detected in mutant strain. The above results indicated that the mutant strain was successfully constructed. HPLC-UV analysis showed that bafilomycins production were abolished in the ΔbafB-E mutant as compared with wild type NA4. This study suggested that the synthetic pathway encoded by bafB-F in S. cavourensis NA4 is the only source of methoxymalonyl-ACP biosynthesis. This study will provide support for rational construction of bafilomycin high-yielding strain through strengthening precursor application strategy.

关 键 词：Ⅰ型聚酮合酶 延伸单元 甲氧基丙二酰ACP 巴弗洛霉素 ΔbafB-E突变株 

分 类 号：Q939.93[生物学—微生物学]