Diphthamide biosynthesis

描述(由申请人提供)：GM088276的A1竞争性更新是一项多学科合作，旨在研究敌草胺的生物合成，这是一种发生在古生物和真核生物翻译延长因子2上的独特蛋白质翻译后修饰。这种修饰已经知道30多年了，是包括白喉毒素在内的几种细菌毒素的靶标。然而，生物合成和生物功能是 人们对此仍知之甚少。以前的研究表明，生物合成的前两步需要五种蛋白质(Dph1-5)，而最后一步(也是最后一步)没有鉴定出蛋白质。有趣的是，几个生物合成基因的缺失在 肿瘤。在之前的资助下，我们取得了一些重要的发现，包括发现了一种新的自由基SAM酶，用于敌草胺生物合成的第一步，并鉴定了生物合成的最后一步所需的两种新蛋白质。这次更新将建立在这些发现的基础上，以进一步了解敌草胺生物合成的化学和酶学，并首次阐明完整的生物合成途径。这项提议将阐明有趣的铁-S酶化学，这将极大地扩展铁-S酶的化学范围和机理理解。对敌草胺生物合成的了解将有助于理解敌草胺的生物学功能以及为什么敌草胺生物合成基因缺失促进肿瘤的发生。

DESCRIPTION (provided by applicant): This A1 competitive renewal of GM088276 is a multidisciplinary collaboration to investigate the biosynthesis of diphthamide, a unique protein posttranslational modification that occurs on archaeal and eukaryotic translation elongation factor 2. This modification has been known for over 30 years and is the target of several bacterial toxins, including diphtheria toxin. However, the biosynthesis and biological function are still poorly understood. Previous studies by others suggest that there are five proteins (Dph1-5) required for the first two steps of the biosynthesis, while no proteins were identified for the thid (and last) amidation step. Interestingly, deletion of several of the biosynthesis genes is found in tumors. With previous grant support, we made a number of important findings, including the discovery of a novel radical SAM enzyme for the first step of diphthamide biosynthesis and the identification of two new proteins required for the last step of the biosynthesis. This renewal wil build on these finding to further understand the chemistry and enzymology of diphthamide biosynthesis and to elucidate the complete biosynthetic pathway for the first time. The interesting Fe-S enzyme chemistry that will be elucidated in this proposal will significantly expand the chemistry scope and mechanistic understanding of Fe-S enzymes. The understanding of diphthamide biosynthesis will help to understand the biological functions of diphthamide and why deletion of diphthamide biosynthesis genes promotes tumorigenesis.