Diphthamide biosynthesis

描述（由申请人提供）：GM 088276的A1竞争性更新是一项多学科合作，旨在研究对苯二甲酰胺的生物合成，对苯二甲酰胺是一种发生在古细菌和真核生物翻译延伸因子2上的独特蛋白质翻译后修饰。这种修饰已经知道了30多年，并且是包括白喉毒素在内的几种细菌毒素的目标。然而，生物合成和生物学功能是 仍然知之甚少。其他人先前的研究表明，生物合成的前两个步骤需要五种蛋白质（Dph 1 -5），而第三个（也是最后一个）酰胺化步骤没有鉴定出蛋白质。有趣的是，发现在大肠杆菌中缺失了几个生物合成基因。 肿瘤的在以前的资助下，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，用于联苯二酰胺生物合成的第一步，并鉴定了生物合成最后一步所需的两种新蛋白质。这一更新将建立在这些发现的基础上，以进一步了解化学和酶学的双苯二甲酰胺生物合成，并阐明完整的生物合成途径的第一次。在本提案中阐明的有趣的Fe-S酶化学将显着扩展Fe-S酶的化学范围和机理理解。对联苯二甲酰胺生物合成的了解将有助于理解联苯二甲酰胺的生物学功能以及为什么联苯二甲酰胺生物合成基因的缺失会促进肿瘤的发生。

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.