Evolution of plant secondary metabolism: The promiscuity of short chain dehydrogenases/reductases involved in tropinone reduction

植物次生代谢的进化：参与托品酮还原的短链脱氢酶/还原酶的混杂

• 批准号: 166283962
• 负责人: Privatdozent Dr. Wolfgang Brandt
• 金额: --
• 依托单位: Abteilung Natur- und Wirkstoffchemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2010
• 资助国家: 德国
• 起止时间: 2009-12-31 至 2013-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/166283962?language=en
• 关键词: Evolution; plant; secondary; metabolism; promiscuity

The goal of the project is to study short-chain dehydrogenases/reductases (SDR) for catalytic capacities and evolvability and to gain information on their native function. Tropinone reduction is a step in tropane alkaloid biosynthesis performed by SDR. A tropinone reductase from tropane alkaloid-forming Cochlearia officinalis (Brassicaceae) has 16 orthologs in A. thaliana annotated as “putative tropine reductase” due to sequence identity of >50%. However, A. thaliana does not accumulate tropane alkaloids. Numerous tropinone reductaselike genes conserved in other Brassicaceae genomes suggest that those SDR form a reservoir for the evolution of secondary product enzymes. We will investigate catalytic characteristics of tropinone reductase-like SDR from Brassicaceae that form or do not form tropane alkaloids. SDR enzyme models serve for ligand identification by in silico pharmacophore search and comparison with known Brassicaceae metabolites. Promising ligands will be tested in vitro. Thus, metabolic flexibility and catalytic redundancy are defined, and hints for the native function of the SDR can be expected. Tropanes are tested as substrates to distinguish tropane-accepting SDR and residues decisive for tropinone binding. SDR are mutated to achieve or enhance tropinone reduction and test model-guided in vitro enzyme evolution.

该项目的目标是研究短链脱氢酶/还原酶（SDR）的催化能力和进化能力，并获得有关其天然功能的信息。托品酮还原是托品烷生物碱生物合成中由SDR进行的步骤。产托品烷生物碱的咖啡豆中的托品酮还原酶在A. thaliana的氨基酸序列由于> 50%的序列同一性而被注释为“推定的托品还原酶”。然而，A.拟南芥不积累托烷生物碱。许多托品酮还原酶样基因保守在其他拟南芥基因组中表明，这些SDR形成了一个水库的进化的次生产物酶。我们将调查的托品酮还原酶样SDR的催化特性，形成或不形成托品烷生物碱。SDR酶模型通过计算机模拟药效团搜索和与已知的拟南芥代谢物的比较用于配体鉴定。有希望的配体将在体外进行测试。因此，代谢的灵活性和催化冗余的定义，并暗示SDR的天然功能可以预期。将托烷作为底物进行测试，以区分托烷接受SDR和对托品酮结合具有决定性的残基。突变SDR以实现或增强托品酮减少和测试模型引导的体外酶进化。