BIOSYNTHESIS OF ANTHRACYCLINE ANTIBIOTICS

该提案要求支持分子遗传学研究， 链霉菌形成蒽环类抗生素的生物化学。 的 一种新抗肿瘤蒽环类代谢物的生物合成 淡蓝链霉菌ETH对四环霉素C（7）的体内活性 22794将作为所有蒽环类抗生素形成的模型进行研究 抗生素 实验方法将首先涉及构建 一组在四环素中具有点阻断的细菌突变体 通过处理S.具有物理和 化学诱变剂 这些突变体将被用作宿主， 用鸟枪基因中来自亲本生物体的DNA片段进行转化 克隆实验，使用已知功能的链霉菌克隆载体 in S.灰绿色的 基因和生化互补的封锁 突变体在转化后产生7的能力将通过 恢复抗生素形成的生物测定;化学和物理的 方法将用于确认这一点以及相关的 内切酶 用高拷贝数载体进行基因扩增，或使用 将探索更高水平的表达载体作为增加 四环素C的酶和中间体的生产 生物合成 辅助工作将完成，以探测蜂窝位置 的抗生素生产基因，以及表达的可能性 这些基因在其他细菌物种中。 研究结果 该计划将提供一种新的方法来研究生物合成的 抗生素，并将建立一般参数所需的增加 生产有价值的抗生素。

This proposal requests support of a study of the molecular genetics and biochemistry of anthracycline antibiotic formation by streptomycetes. The biosynthesis of a new anthracycline metabolite with potent antitumor activity in vivo, tetracenomycin C (7), by Streptomyces glaucescens ETH 22794 will be studied as a model of the formation of all anthracycline antibiotics. The experimental approach will first involve construction of a set of bacterial mutants having point blocks in the tetracenomycin biosynthetic pathway by treatment of S. glaucescens with physical and chemical mutagens. These mutants will then be used as hosts for transformation with DNA segments from the parent organism in shotgun gene cloning experiments, using Streptomyces cloning vectors known to function in S. glaucescens. Genetic and biochemical complementation of the blocked mutants' ability to produce 7 after transformation will be assessed by bioassay for the restoration of antibiotic formation; chemical and physical methods will be used to affirm this and the presence of the associated enzymes. Gene amplification with high copy number vectors, or the use of high level expression vectors will be explored as a means for increasing the production of the enzymes and intermediates of tetracenomycin C biosynthesis. Subsidiary work will be done to probe the cellular location of the antibiotic production genes, and the possibility for expression of these genes in other bacterial species. The results of the research program will exemplify a new approach to research on the biosynthesis of antibiotics, and will establish general parameters necessary for increasing the production of valuable antibiotics.