BIOSYNTHESIS OF ANTHRACYCLINE ANTIBIOTICS

蒽环类抗生素的生物合成

• 批准号: 2088942
• 负责人: CHARLES R HUTCHINSON
• 金额: $ 21.45万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-09-01 至 1997-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2088942
• 关键词: Escherichia coli; Streptomyces; X ray crystallography; active sites; acyltransferase; anthracyclines; antineoplastic antibiotics; antiviral agents; bacterial genetics; biotechnology; cell growth regulation; enzyme complex; enzyme mechanism; enzyme reconstitution; enzyme structure; fatty acid metabolism; fatty acid synthase; fluorescence spectrometry; gene expression; gene mutation; genetic manipulation; genetic regulation; genetic transcription; ketones; microorganism metabolism; nuclear magnetic resonance spectroscopy; nucleic acid sequence; radiotracer; tissue /cell culture; western blottings

I propose to continue a study of the biochemistry and genetics of the production of tetracenomycin C (Tcm C), a metabolite of Streptomyces glaucescens that is classified chemically as an anthracycline and has moderate antitumor properties, as a model of secondary metabolism. Information about the biosynthesis of polyketides, like Tcm C, can be used to increase the production of known drugs. In attempts to make new drugs by genetic engineering, which is an attractive prospect for anthracyclines because some of them have clinically valuable antitumor and notable anti- HIV activity, the knowledge that the polyketides represent hundreds of different structural types suggests that numerous perturbations of this biochemistry are possible. This could make the search for new drugs by genetic engineering especially fruitful. Antibiotic production is a characteristic of Streptomyces that has attracted considerable attention. Probing its genetics is likely to uncover significantly new information because secondary metabolism, a unique characteristic of the slow growth (stationary) phase of laboratory cultures, is quite unlike the primary metabolism of rapidly growing cells that has been the focal point of most studies of prokaryotic genetics. Moreover, the value of using the PKS genes and genes that regulate antibiotic production to construct strains that overproduce secondary metabolites has been demonstrated; thus, information about the regulatory mechanisms should have wide-spread utility in biotechnology. Our goals for the next five years, listed in the priority in which they will be pursued, are as follows. 1. Enzymology of the tcm polyketide synthase (PKS). We will study the enzymology of polyketide metabolism by (i) determining the properties of the enzymes produced by the tcm PKS genes, (ii) using these enzymes to develop a system for the production of Tcm F2 nd Tcm F1 (or D3) in vitro, and then (iii) investigating the effect of substituting the components of other PKS's for the different enzymes of the tcm PKS. New metabolites may be produced in this work. 2. Regulation of the expression of the tcm genes. We will investigate the genetics of Tcm C production by (i) elucidating the transcriptional organization of the tcmIII/VI/Ia/II, tcmVII, and tcmAR genes, (ii) determining whether expression of the tcm PKS, tcmVII and tcmA genes is controlled by the tcmR gene or vice-versa, and (iii) searching for genes or physiological factors that control expression of the tcmIII/VI/Ia/II and tcmAR genes. This will provide insight into the regulation of secondary metabolism. 3. Properties of the fatty acid synthase (FAS) genes. We will determine the effect of deleting the Orf1 gene of the S. glaucescens ACP-II region on cell growth and development, and the production of the Tcm C metabolites. This work will reveal possible interactions between polyketide and fatty acid metabolism and whether the Orf1 gene is part of the FAS.

我建议继续研究的生物化学和遗传学的 链霉菌代谢产物四环霉素C的生产 在化学上被分类为蒽环类， 中度抗肿瘤特性，作为次级代谢的模型。 关于聚酮化合物的生物合成的信息，如Tcm C，可用于 以增加已知药物的产量。 为了制造新药 这是蒽环类抗生素的一个诱人的前景 因为其中一些化合物具有临床上有价值的抗肿瘤作用， 艾滋病毒活性，聚酮化合物代表数百种 不同的结构类型表明， 生物化学是可能的。 这可能会使新药的研究， 基因工程尤其富有成效。 抗生素生产是链霉菌的一个特征， 引起了相当大的关注。 探索它的基因很可能 揭示了重要的新信息，因为次级代谢， 实验室缓慢生长（稳定）期的独特特征 与快速生长的细胞的初级代谢完全不同， 这是大多数原核生物遗传学研究的焦点。 此外，使用PKS基因和调节PKS的基因的价值在于， 抗生素生产，以构建过量产生次级 代谢物已被证明;因此，有关监管的信息 这些机制应在生物技术中具有广泛的用途。 我们未来五年的目标，列在优先事项中， 将遵循如下。 1.中药聚酮的酶学研究 合成酶（PKS）。 我们将研究聚酮代谢的酶学， (i)确定由中药PKS产生的酶的性质 基因，（ii）使用这些酶来开发用于生产 Tcm F2和Tcm F1（或D3）体外培养，然后（iii）研究 用其他PKS的成分代替不同的酶， 中药PKS。 在这项工作中可能会产生新的代谢产物。 2.调控 中医药基因的表达。 我们将研究 通过（i）阐明Tcm C的转录组织， tcm III/VI/Ia/II、tcm VII和tcm AR基因，（ii）确定是否 tcm PKS、tcm VII和tcm A基因的表达受tcmR控制， 基因或反之亦然，以及（iii）搜索基因或生理因子 其控制tcm III/VI/Ia/II和tcm AR基因的表达。 这将 提供对次级代谢调节的深入了解。 3.性能 脂肪酸合成酶（FAS）基因。 我们将确定 缺失S.灰毛苔ACP-II区对细胞生长的影响 和开发，以及Tcm C代谢产物的产生。 这项工作 将揭示聚酮化合物和脂肪酸之间可能的相互作用 代谢和Orf 1基因是否是FAS的一部分。