基因毒性抗生素阿嗪霉素B产生菌自抗性及交联DNA损伤修复机制的研究

基因毒性抗肿瘤抗生素阿嗪霉素B的化学性质非常活泼，很容易与鸟嘌呤G发生烷基化反应引起DNA交联。其产生菌链霉菌Streptomyces sahachiroi的基因组中富含有能够被阿嗪霉素B攻击的靶标位点，由此引发了我们对产生菌抵抗DNA交联损伤机制的研究兴趣。前期研究从药物的抗性基因入手，发现了两个新型的DNA损伤修复蛋白，转葡糖基酶AlkZ和结构特异性核酸内切酶AziN。本项目拟综合运用遗传学、分子生物学、生物化学的研究方法，对该菌中另外三个潜在的抗性因子AziE、AziF和Orf8展开功能研究，详细阐明它们对阿嗪霉素B的抗性作用机理，进一步完善阿嗪霉素B产生菌的自抗性系统。同时通过寻找能够协助抗性因子AlkZ和AziN修复交联DNA的下游功能蛋白，在分子水平上深入剖析这两种DNA损伤修复途径的完整过程、作用机制以及它们之间的相关性，建立产生菌综合利用不同DNA修复途径来抵御药物分子的作用模型。

The active genotoxic antibiotic azinomycin B has prominent in vivo antitumor activity and in vitro cytotoxicity by alkylating guanine nucleosides in duplex DNA and inducing interstrand crosslinks. On the basis of an average G+C content of 75%, there may be a large number of potentially susceptible target sites within the genome of the azinomycin B producing strain Streptomyces sahachiroi. Therefore, the mechanism of self-protection to prevent from DNA cross-linking damage in the producer aroused our research interest. Through searching for the drug resistance genes, two novel proteins including the DNA glycosylase AlkZ and the structure-specific endonuclease AziN were found to be implicated in the repair of azinomycin B-mediated DNA interstrand crosslinks by our previous studies. In this project, functional study of three other potential resistance factors AziE, AziF and Orf8 by using genetics, molecular biology and biochemistry methods will disclose their resistance mechanisms and comprehensively elucidate the self-resistance system in S. sahachiroi. Meanwhile, discovery of more DNA repair factors incorporated with AlkZ and AziN will not only contribute to clearly elucidate the complete process of two cross-linked DNA repair strategies and their correlation, but also set a basis to establish a model of the comprehensive utilization of different DNA repair pathways in the producer to protect itself from the lethal effects of drug.