Approach to a enedyine biosynthesis tool box through metagenomic and targeted metabolomic analyses of actinomycetes

通过放线菌宏基因组和靶向代谢组分析建立烯啶生物合成工具箱

• 批准号: 276972917
• 负责人: Dr. Ivana Crnovcic
• 金额: --
• 依托单位: Department of Chemistry
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2016-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/276972917?language=en
• 关键词: Approach; enedyine; biosynthesis; tool; box

Enediynes are a class of potent antitumor agents produced by soil and marine microorganisms. Structurally they are characterized by a core unit consisting of two acetylenic groups conjugated to a double bond or incipient double bond within a 9- or 10-membered ring. The enediynes possess anticancer and antibiotic activity based on the presence of their enediyne core whereas the peripheral chemistry found around the cores of the different enediynes plays only a role in the fine tuning of that activity. Enediynes produce in the DNA inter-strand DNA crosslinks (ICLs) and can lead to site-specific single stranded or double stranded breaks (DSBs). Comparison of the known enediyne biosynthetic gene clusters has revealed that both, 9- and 10-membered enediynes, share a conserved five-gene cassette, the enediyne PKS cassette, which encodes the biosynthesis of the pentadecaheptaene backbone of the enediyne core structure. The conservation of the central core of the pentadecaheptaene is disturbed by the peripheral parts which lead to a high structural diversity of the various enediynes. Since the enediynes have extreme cytotoxic activity as pure compounds they have only limited use as clinical drugs. However, when formulated into various polymer-based delivery systems or as anticancer antibody-drug conjugates (ADCs) they have shown great clinical success and are very promising for future developments in anticancer therapy. Therefore the biosynthetic development of new enediyne compounds with still better performance is of interest. The group of Prof. Shen has done a genome survey of 3,200 strains of their Actinomycetale collection and identified 89 strains that harbor the enediyne PKS gene cassettes. In detail, two strains, named CB02115 and CB100675 from this strain collection we will chose as models to investigate novel structural features such as e. g. specific groupings or functional groups in their biosynthesis. CB02115 encodes a gene cluster that resembles the C-1027 gene cluster from Streptomyces globisporus, while CB100675 encodes a gene cluster that shows similarity to the calicheamicin (CAL) gene cluster from Micromonospora echinospora. To address the structure and the biosynthetic features of the enediynes encoded by the enediyne gene cluster in strain CB02115 and strain CB100675 we will determine the structures of the new enediynes and analyse promising genes with apparent novel biosynthetic function in enediyne biosynthesis. Moreover, in view of the similarity of the two new enediyne gene clusters with already known clusters we will express the gene constructs in a genetic background actively synthesizing conventional enediynes. For the most successful genes in either CB02115 or CB100675 we will in vitro characterize its gene product by expressing the gene in E. coli and testing the enzyme with a suitable model substrate in vitro.

烯二炔是一类由土壤和海洋微生物产生的有效抗肿瘤剂。在结构上，它们的特征在于由两个炔基组成的核心单元，所述炔基与9元或10元环内的双键或初始双键缀合。基于其烯二炔核心的存在，烯二炔具有抗癌和抗生素活性，而在不同烯二炔核心周围发现的外围化学仅在该活性的微调中起作用。烯二炔在DNA中产生链间DNA交联（ICL），并可导致位点特异性单链或双链断裂（DSB）。已知的烯二炔生物合成基因簇的比较已经揭示，9元和10元烯二炔都共享保守的五基因盒，烯二炔PKS盒，其编码烯二炔核心结构的十五碳七烯骨架的生物合成。十五碳七烯的中心核的保守性受到外围部分的干扰，这导致各种烯二炔的高度结构多样性。由于烯二炔作为纯化合物具有极端的细胞毒性活性，因此它们作为临床药物的用途有限。然而，当配制成各种基于聚合物的递送系统或作为抗癌抗体-药物缀合物（ADC）时，它们已经显示出巨大的临床成功，并且对于抗癌疗法的未来发展非常有希望。因此，具有更好性能的新烯二炔化合物的生物合成开发是令人感兴趣的。沈教授的研究小组对放线菌的3,200株菌株进行了基因组调查，并鉴定出89株含有烯二炔PKS基因盒的菌株。详细地说，我们将从该菌株收集物中选择两个菌株，命名为CB 02115和CB 100675作为模型来研究新的结构特征，例如e. G.在其生物合成中的特定分组或功能组。CB 02115编码的基因簇类似于来自球孢链霉菌的C-1027基因簇，而CB 100675编码的基因簇显示出与来自棘孢小单孢菌的加利车霉素（CAL）基因簇的相似性。为了解决菌株CB 02115和菌株CB 100675中烯二炔基因簇编码的烯二炔的结构和生物合成特征，我们将确定新的烯二炔的结构，并分析在烯二炔生物合成中具有明显的新生物合成功能的有前途的基因。此外，鉴于两个新的烯二炔基因簇与已知簇的相似性，我们将在积极合成常规烯二炔的遗传背景中表达基因构建体。对于CB 02115或CB 100675中最成功的基因，我们将通过在E.大肠杆菌，并在体外用合适的模型底物测试酶。

项目成果

Strain Prioritization and Genome Mining for Enediyne Natural Products.

• DOI: 10.1128/mbio.02104-16
• 发表时间: 2016-12-20
• 期刊: mBio
• 影响因子: 6.4
• 作者: Yan X;Ge H;Huang T;Hindra;Yang D;Teng Q;Crnovčić I;Li X;Rudolf JD;Lohman JR;Gansemans Y;Zhu X;Huang Y;Zhao LX;Jiang Y;Van Nieuwerburgh F;Rader C;Duan Y;Shen B
• 通讯作者: Shen B

Engineered production of cancer targeting peptide (CTP)-containing C-1027 in Streptomyces globisporus and biological evaluation.

靶向肽（CTP）的工程生产 - 全球链霉菌和生物学评估中含有C-1027。

• DOI: 10.1016/j.bmc.2016.04.017
• 发表时间: 2016-09-01
• 期刊: Bioorganic & medicinal chemistry
• 影响因子: 3.5
• 作者: Li W;Li X;Huang T;Teng Q;Crnovcic I;Rader C;Shen B
• 通讯作者: Shen B

Discovery of Alternative Producers of the Enediyne Antitumor Antibiotic C-1027 with High Titers.

• DOI: 10.1021/acs.jnatprod.7b01013
• 发表时间: 2018-03-23
• 期刊: Journal of natural products
• 影响因子: 5.1
• 作者: Yan X;Hindra;Ge H;Yang D;Huang T;Crnovcic I;Chang CY;Fang SM;Annaval T;Zhu X;Huang Y;Zhao LX;Jiang Y;Duan Y;Shen B
• 通讯作者: Shen B