Charakterisierung eines neu entdeckten mitochondrialen molybdäncofaktorabhängigen Enzymsystems

新发现的线粒体钼辅因子依赖性酶系统的表征

• 批准号: 111328339
• 负责人: Professor Dr. Bernd Clement
• 金额: --
• 依托单位: Pharmazeutisches Institut
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2009
• 资助国家: 德国
• 起止时间: 2008-12-31 至 2011-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/111328339?language=en
• 关键词: Charakterisierung; eines; neu; entdeckten; mitochondrialen

The "mitochondrial amidoxime reducing component" (mARC) is a molybdenum-containing enzyme, that was discovered in our labs in 2006. Besides mARC, only three other molybdoenzymes are known in humans. This enzyme is able to reduce N-hydroxylated compounds together with cytochrome b5 (cyt b5) and NADH cytochrome b5 reductase (b5 Red). Almost all eukaryotic genomes encode for two mARC enzymes (mARC1 and mARC2), which are very similar in respect to their amino acid sequence. In addition, both mARC enzymes are able to reduce N-hydroxylated compounds together with cyt b5 and b5 Red.Based on our previous work and publications the enzyme "mARC" is has been successfully established in the scientific world. Thus, the designation "mARC" proposed by us was adopted by the HUGO Gene Nomenclature Committee (HGNC) in October 2011. The molybdenumcofactor-dependend enzyme system was originally discovered during investigations of N-reductive drug metabolism. This involvement in drug metabolism is now well investigated, our primary objective is therefore the exploration of the physiological function of the enzyme system. Our proposed projects are primarily based on our previous in vitro studies using isolated recombinant enzymes or subcellular organelles. In order to better assess the physiological relevance of this enzyme system our future studies should be carried out mainly with cellular systems and also using our mARC-knockout mouse model. In particular the following aspects should be considered: (i) Are one or both mARC enzymes involved in the N-reductive pathway? (ii) Is the enzyme system a cellular protection mechanism that prevents incorporation of mutagenic base analogues into the nucleic acids of the cell? (iii) Does the enzyme system modulates the metabolism of L-arginine and NO and is therefore involved in diabetes/diabetic nephropathy? (iv) Is the subcellular localization in mitochondria and peroxisomes a hint of a potential connection of mARC with the complex metabolism of reactive oxygen species? As the mARC-containing enzyme system plays a central role in the metabolism of drugs with N-hydroxylated functional groups, our additional aims are: (v) The characterization of the reductive detoxification reactions of N-hydroxylated aniline derivatives. In this context genetic polymorphisms of the N-reductive enzyme system should be taken into account. (vi ) The investigation of quantitative structure-activity relationships between the chemical structure of the substrates and the reductase activity. This information could be later applied to the rational development of so-called "amidoxime" prodrugs. Our planned work to (vii) characterize biochemical-spectroscopically both the active center and functionally important protein sequence motifs, and (viii) the elucidation of the atomic structure of mARC enzymes will contribute to the general understanding of this novel class of enzymes.

“线粒体偕胺肟还原组分”（mARC）是一种含氘的酶，于2006年在我们的实验室中发现。除了mARC，人类中已知的其他三种酶只有三种。这种酶能够与细胞色素b5（cyt b5）和NADH细胞色素b5还原酶（b5 Red）一起还原N-羟基化化合物。几乎所有的真核生物基因组都编码两种mARC酶（mARC 1和mARC 2），它们的氨基酸序列非常相似。此外，这两种mARC酶都能够与cyt b5和b5 Red一起还原N-羟基化化合物。基于我们以前的工作和出版物，酶"mARC"已在科学界成功建立。因此，我们提出的名称“mARC”于2011年10月被HUGO基因命名委员会（HGNC）采纳。辅酶依赖酶系统最初是在研究N-还原性药物代谢过程中发现的。这种参与药物代谢的研究现在很好，因此我们的主要目标是探索酶系统的生理功能。我们提出的项目主要是基于我们以前的体外研究，使用分离的重组酶或亚细胞器。为了更好地评估这种酶系统的生理相关性，我们未来的研究应该主要用细胞系统进行，也可以使用我们的mARC敲除小鼠模型。具体而言，应考虑以下方面：（i）一种或两种mARC酶是否参与N-还原途径？(ii)酶系统是否是一种细胞保护机制，防止诱变碱基类似物掺入细胞核酸中？(iii)酶系统是否调节L-精氨酸和NO的代谢，从而参与糖尿病/糖尿病肾病？(iv)线粒体和过氧化物酶体的亚细胞定位是否暗示mARC与活性氧的复杂代谢存在潜在联系？由于含MARC的酶系统在具有N-羟基化官能团的药物的代谢中起着核心作用，我们的额外目标是：（V）N-羟基化苯胺衍生物的还原解毒反应的表征。在这种情况下，应考虑到N-还原酶系统的遗传多态性。(vi研究底物的化学结构与还原酶活性之间的定量构效关系。这一信息可以在以后应用于所谓的"偕胺肟"前药的合理开发。我们计划的工作，（七）生化光谱表征的活性中心和功能重要的蛋白质序列基序，和（八）阐明的原子结构的mARC酶将有助于这类新的酶的一般理解。