Molecular mechanisms and roles of the mammalian 2-oxoglutarate-dependent oxygenases AlkBH1 and AlkBH7

基因和蛋白质的活性独立于DNA遗传密码，通过写入、读取和擦除特定的表观遗传标记，由表观遗传机制控制。这些标记包括组蛋白、DNA和RNA与甲基的共价修饰。这些甲基在许多细胞过程中起着至关重要的作用。负责这些表观遗传标记的空间和时间维持是由所谓的“写入者”和“擦除者”酶执行的。其中，依赖于Fe~(2+)和2-氧戊二酸(2OG；=α-酮戊二酸)的加氧酶家族中的加氧酶尤为重要。到目前为止，在所有的生命王国中，大约有70名Fe(II)/2OG-家族成员是已知的。这些酶使用辅基2OG与亚铁结合并激活分子氧，以催化分子氧转移到甲基上，从而产生稳定的羟化产物(=羟基酶)或通过氧化去甲基化(=去甲基酶)使底物去甲基化。这项建议涉及的亚家族是AlkB同系物(AlkBHs)，它们存在于各种生物中，如大肠杆菌、果蝇、裂殖酵母、秀丽线虫和哺乳动物。目前在哺乳动物中有九种AlkB同系物：AlkBH1-8和脂肪质量和肥胖相关基因产物(FTO)。由于它们在序列、结构元素和辅助结构域上的差异，导致它们在底物和生物学作用上存在着巨大的多样性。研究表明，对于单个哺乳动物AlkBH，其功能丧失及其异常表达和活性可能与各种疾病表型有关，如肥胖、对炎症严重敏感、多发性畸形、不孕不育和癌症，这强调了这些酶的重要性。然而，许多人类碱性血球蛋白的分子功能、观察到的表型和底物范围之间的直接联系仍然不清楚。在这里提出的研究项目中，我们的目的是阐明最难以捉摸的人类碱性氢化氢的分子机制。我们想了解他们的底物和相互作用网络。我们的重点将是人类的AlkBH1和ALKBH7蛋白。所采用的方法包括细胞生物学、质谱学和结构生物学。

Independent from the DNA genetic code, gene and protein activity is controlled by epigenetic mechanisms through writing, reading and erasing specific epigenetic marks. These marks include the covalent modifications of histones, DNA and RNA with methyl groups. These methyl groups play an essential role in many cellular processes. Responsible for the spacial and temporal maintenance of these epigenetic marks is carried out by so called "writer" and "eraser" enzymes. Here enzymes belonging to the Fe2+- and 2-oxoglutarate (2OG; = α-ketoglutarate) dependent oxygenase family are particular prominent actors. About 70 members of the Fe(II)/2OG-family across all kingdoms of life are known to date. These enzymes use the cosubstrate 2OG to bind to ferrous iron and to activate molecular oxygen in order to catalyse the transfer of the molecular oxygen onto a methyl group, resulting either in a stable hydroxylated product (=hydroxylases) or demethylation of the substrate via oxidative demethylation (=demethylases). The subfamily this proposal addresses are the AlkB-homologs (AlkBHs), which are found in various organisms such as Escherichia coli, Drosophila melanogaster, Schizosaccharomyces pombe, Cenorhabditis elegans and mammals.In mammals there are nine AlkB-homologs known by now: AlkBH1-8 and the fat mass- and obesity-associated gene product (FTO). The differences in sequence, structural elements and accessory domains account for their remarkably large diversity in substrates and biological roles. For individual mammalian AlkBHs it was shown, that loss-off function as well as their aberrant expression and activities can be linked to various diseases phenotypes, such as obesity, severe sensitivity to inflammation, multiple malformations, infertility and cancer, which emphasizes the importance of these enzymes. However, a direct link between the molecular functions, observed phenotype and the substrate scope for many of the human AlkBHs is still unclear. In the here proposed research project our aim is to elucidate the molecular mechanism of the most elusive human AlkBHs. We want to learn about their substrates and interaction network. Our focus will be the human AlkBH1 and AlkBH7 proteins. The employed methods involve cell biology, mass spectrometry and structural biology.