DAUNORUBICIN DPSH GENE ON CYCLIZATION PATTERN FOR TYPE II POLYKETIDE SYNTHASE

II 型聚酮合酶环化模式中的柔红霉素 DPSH 基因

• 批准号: 6309123
• 负责人: CHARLES R HUTCHINSON
• 金额: $ 0.75万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-15 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6309123
• 关键词: bacteria; bioengineering /biomedical engineering; biological products; biomaterials; biomedical resource; biotechnology; enzymes; nuclear magnetic resonance spectroscopy

Production of aklanoic acid, an early intermediate of daunorubicin (dnr) biosynthesis in Streptomyces peucetius, by hybrid type II polyletide synthase (PKS) genes was studied as a function of the order and type of genes cloned in a plasmid vector and expressed from the ermE* promoter. Compared with expression of the daunorubicin polyketide synthase (dps) genes in the normal order from their native promoter, the dnrG monooxygenase, dpsA and dpsB ketoreductase and dpsF cyclase genes, when cloned in this order in Streptomyces lividans, produced aklanoic acid and SEK-43. The latter compound is made from acetyl-coenzyme A instead of propionyl-coenzyme A as the starter unit and does not have the characteristic fused ring aromatic system of aklanoic acid. Replacement of either dpsG with its tcmM homolog or dnrG with the Streptomyces glaucescens tcmJ PKS gene resulted in little or no aklanoic acid production. In contrast, addition of the dpsH gene to the tcmJ, dpsA, dpsB, tcmM, dpsE and dpsF gene cassette restored cyclization pattern of aklanoic acid. The reason for the ability of dpsH to override the negative effect of tcmM and restore both the proper choice of starter unit and cyclization pattern is unknown, but overall results indicate that iterative type II PKSs can function aberrantly, perhaps as a function of abnormal protein stoichiometries or interactions. Currently, we are investigating structures of compounds when these gene cassettes are expressed in other mutants.

柔红霉素的早期中间体--链烷酸的生产 (dnr)在Streptomyces peucetius中的生物合成，通过杂交II型 多肽合成酶（PKS）基因被研究作为顺序的功能 和克隆在质粒载体中并从表达载体表达的基因的类型。 ermE* 启动子。 与柔红霉素的表达相比， 聚酮合酶（DPS）基因以正常顺序从其天然的 启动子、dnrG单加氧酶、dpsA和dpsB酮还原酶和dpsF 环化酶基因，当在变铅青链霉菌中按此顺序克隆时， 产生链烷酸和SEK-43。 后一种化合物由 乙酰辅酶A代替丙酰辅酶A作为起始单元 并且不具有特征性的稠环芳族体系， 链烷酸 用其tcmM同系物替换dpsG，或 dnrG与灰绿链霉菌tcmJ PKS基因的融合导致 很少或没有链烷酸产生。 相比之下，添加 dpsH基因与tcmJ、dpsA、dpsB、tcmM、dpsE和dpsF基因盒连接 链烷酸的环化模式恢复。 的原因 dpsH克服tcmM的负面作用并恢复 选择合适的起始剂和环化方式， 未知，但总体结果表明，迭代II型PKS可以 功能异常，可能是异常蛋白质的功能 化学计量或相互作用。 目前，我们正在调查 当这些基因盒在细胞中表达时， 其他变种人