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一种新型酰基辅酶A羧化酶的发现、研究及在聚酮骨架改造中的应用
Polyketides Scaffold Engineering Based on Extender Units Synthesis by a Permissive Acyl-CoA Carboxylase
【作者】 张俊;
【导师】 瞿旭东;
【作者基本信息】 武汉大学 , 药学, 2021, 博士
【摘要】 聚酮类天然产物数量庞大,结构复杂且生物活性良好,被广泛应用于医药和农业,其药理活性与分子结构密不可分。基于聚酮天然产物的生物合成逻辑,科学家可以便捷的操纵聚酮生物合成途径并理性改造聚酮分子。在聚酮天然产物结构多样性的所有成因中,合成砌块延伸单元是聚酮碳骨架多样化最重要的来源。因此,基于延伸单元的碳骨架重构被认为是最有效且彻底的聚酮分子改造途径。生物体中,聚酮合酶烷基丙二酸单酰辅酶A类型的延伸单元主要通过三种生物合成途径获得,其中,酰基辅酶A羧化酶途径一步合成延伸单元,不依赖双键底物且不产生还原型副产物,具有天然的优势。本研究从聚酮天然产物碳骨架的改造思路出发,利用延伸单元为切入点,基于已有的链霉菌Streptomyces armeniacus DSM 43125中活性聚酮分子armeniaspirol生物合成研究结果,发现了一例新型酰基辅酶A羧化酶β亚基Arm13。进一步对Arm13进行探究,发现该β亚基可以与不同来源的α亚基与ε亚基组装成一个具有高度催化活性的羧化酶复合体;体外催化研究结果表明该羧化酶复合体可以识别各种链长(C4-C10)以及末端烯基、炔基、卤素、苯环、吲哚等各种官能团的酰基辅酶A底物,并高效转化为相应的丙二酸单酰辅酶A类型延伸单元。基于Arm13强大的延伸单元合成能力,本研究通过对armeniaspirol产生菌S.armeniacus DSM43125喂养简单的羧酸甲酯前体,成功向armeniaspirol聚酮骨架C6位引入了烯基、炔基活性官能团以及芳香性的苯环,证实Arm13在聚酮碳骨架编辑方面的应用潜力。本研究成果不仅促进了对新型中链酰基辅酶A羧化酶β亚基的亚基适配性与底物谱范围认知,也为基于延伸单元的聚酮骨架合理设计与改造提供了强有力的工具酶。
【Abstract】 Polyketides are a large class of structurally diverse natural products possessing various pharmaceutically important activities.In spite of their large structural and functional variety,polyketides are all assembled from simple acyl-and alkylmalonylthioester building blocks through successive decarboxylate Claisen-condensation reactions catalyzed by polyketide synthases.This conserved biosynthetic mechanism of polyketide synthases facilitates engineering efforts to manipulate the assembling process for rational modification of their carbon scaffolds.The structural diversity of polyketides can be attributed to a variety of factors,from which,extender units were thought to contribute mostly for polyketide diversity,thus one particularly important engineering strategy is the use of different extender units(S)-alkylmalonyl-Co A during polyketide synthesis.(S)-alkylmalonyl-Co As can be formed by three ways,among them,acyl-Co A carboxylases pathway comprised of one step and independent of α,β-double bond substrates.Furthermore,acyl-Co A carboxylases pathway won’t produce reductive sideproducts,thus,it could be more appealing for avoiding such inherent drawbacks in extender units biosynthesis.Here,we report a novel β-subunit Arm13 of acyl-Co A carboxylases involved in biosynthesis of armeniaspirols in Streptomyces armeniacus DSM 43125.Through comprehensive in vivo and in vitro characterization,we established that Arm13 is permissive to nonnative α-and ε-subunits to form a highly promiscuous acyl-Co A carboxylases complex able to accept various functionalized medium chain acyl-Co As and maintain high catalytic efficiency.By applying this biosynthetic machinery in vivo,we introduced a series of structural variations into the carbon scaffolds of armeniaspirols with simple precursors,demonstrating its broad utility in providing extender units for polyketides carbon scaffold editing.These findings not only enrich the knowledge of the medium chain specific acylCo A carboxylases,but also provide a versatile biocatalyst for the manipulation of building blocks and set the stage for the rational design of polyketide biosynthesis.
【Key words】 extender units; polyketide scaffold; Acyl-Co A carboxylase; armeniaspirol;
- 【网络出版投稿人】 武汉大学 【网络出版年期】2025年 04期
- 【分类号】R915