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MenD (SEPHCHC synthase), a thiamine-dependent enzyme with similarity to the Stetterases

MenD（SEPHCC 合酶），一种与 Stetterase 相似的硫胺素依赖性酶

基本信息

批准号：
172064182
负责人：
Professor Dr. Georg Sprenger
金额：
--
依托单位：
依托单位国家：
德国
项目类别：
Research Units
财政年份：
2010
资助国家：
德国
起止时间：
2009-12-31 至 2016-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/172064182?language=en
关键词：
MenD SEPHCHC synthase thiamine dependent

项目摘要

As the only thiamine diphosphate-dependent enzyme, MenD (SEPHCHC synthase) from the bacterial menaquinone biosynthetic pathway, performs an 1,4 addition reaction on a ring structured acceptor compound which neither displays an aldehyde nor a keto structure. Moreover, the working group of Michael Müller has discovered an interesting 1,2 addition on arylaldehydes, thus allowing the synthetic access to novel 2-hydroxy ketones. Besides a-ketoglutarate (a-oxoglutarate), MenD is able to utilize pyruvate or oxaloacetate as donor compounds. The aim of our work in the frame of the project is to analyze the substrate spectrum (both donor and acceptor side) of MenD in more detail and to make the enzyme applicable in preparative syntheses. Based on the 3D structure of the enzyme - which became available recently- we want to cooperate with a modeling group and study the function of all amino acid residues of the enzyme s active site. Special emphasis will be on residues which are involved in substrate binding and/or the catalysis of 1,2 and 1,4 additions. Using site-directed saturation mutagenesis, we will aim for enzyme variants with altered substrate specificities. As well, substrate analogues which inhibit MenD activity may provide interesting lead structures for novel antibiotics as the menaquinone biosynthetic pathway is essential for several pathogenic bacteria, e.g. Mycobacterium tuberculosis.

MenD（SEPHCHC synthase，SEPHCHC合酶）作为唯一的二磷酸硫胺素依赖性酶，在一个既不含醛也不含酮结构的环状受体化合物上进行1，4加成反应。此外，Michael Müller的工作组发现了一个有趣的芳基醛的1，2加成，从而允许合成新的2-羟基酮。除了α-酮戊二酸（α-酮戊二酸），MenD能够利用丙酮酸或草酰乙酸作为供体化合物。我们在该项目框架内工作的目的是更详细地分析MenD的底物光谱（供体和受体侧），并使酶适用于制备合成。基于酶的3D结构-最近可用-我们希望与建模小组合作，研究酶活性位点的所有氨基酸残基的功能。特别强调的是参与底物结合和/或催化1，2和1，4加成的残基。使用定点饱和诱变，我们将针对具有改变的底物特异性的酶变体。同样，抑制MenD活性的底物类似物可以为新型抗生素提供有趣的先导结构，因为甲萘醌生物合成途径对于几种病原菌（例如结核分枝杆菌）是必需的。