Diphthamide biosynthesis

二邻苯二甲酰胺生物合成

• 批准号: 8307724
• 负责人: STEVEN E EALICK
• 金额: $ 30.97万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8307724
• 关键词: ADP ribosylation; Adenosine Diphosphate Ribose; Amino Acid Sequence; Anabolism; Archaea; Bacterial Infections; Bacterial Toxins; Binding; Biochemical; Biological Process; Biology; Cell Death; Cells; Chemistry; Complex; Cytosol; Data; Development; Diphtheria Toxin; Electron Spin Resonance Spectroscopy; Enzymatic Biochemistry; Enzymes; Eukaryota; Eukaryotic Cell; Exotoxins; Genes; Genetic; Goals; Health; Histidine; Human; Imidazole; In Vitro; Iron; Knowledge; Lead; Learning; Link; Literature; Malignant Neoplasms; Malignant neoplasm of ovary; Methods; Methyltransferase; Modification; Mossbauer Spectroscopy; Mutation; Names; Nicotinamide adenine dinucleotide; Nitrogen; Oncogenes; Pathway interactions; Peptide Elongation Factor 2; Peptide Sequence Determination; Post-Translational Protein Processing; Protein Biosynthesis; Proteins; Pseudomonas aeruginosa toxA protein; Reaction; Regulation; Reporting; S-Adenosylmethionine; Structure; Sulfur; Testing; Toxin; Translations; Tumor Suppression; Tumor Suppressor Proteins; Ursidae Family; amidation; base; in vitro Assay; in vivo; insight; interest; malignant breast neoplasm; pathogen; prevent; tumor

DESCRIPTION (provided by applicant): Diphthamide is a post-translationally modified histidine residue found in translation elongation factor 2 (eEF-2) in all eukaryotic cells. This single modification concentrates a lot of interesting chemistry and biology. This modification involves a unique C-C bond formation reaction that links the C2 of the imidazole ring and the 3-amino-3-carboxyl propyl group derived from S-adenosylmethionine (SAM). This C-C bond formation reaction is followed by a trimethylation step and an amidation step to give the final diphthamide modification. During certain bacterial infections, this modified residue (but not the unmodified histidine residue) is specifically recognized by bacterial toxins, including diphtheria toxin and Pseudomonas exotoxin A. These toxins catalyze the transfer of an adenosine diphosphate ribose (ADP-ribose) from nicotinamide adenine dinucleotide (NAD) to one of the nitrogen atoms in the imidazole ring. The ADP-ribosylation by bacterial toxins inhibits the function of eEF-2 and thus protein synthesis, leading to cell death. Based on genetic studies, the first step of the biosynthesis, the formation of the C-C bond, requires four proteins in eukaryotes, Dph1, Dph2, Dph3, and Dph4. However, how these four proteins catalyze the reaction and why so many proteins are involved in one reaction is unknown. Heterozygous deletions of dph1 and dph4 genes are associated with several human cancers, particularly ovarian and breast cancers, raising interesting questions about the biological function of diphthamide and the tumor suppression mechanism of Dph1 and Dph4. In this proposal, we aim to study how Dph1, Dph2, Dph3, and Dph4 catalyze the first step of diphthamide biosynthesis. Understanding the biosynthesis will uncover some very interesting enzymology, as predicted by our preliminary results, and provide insights into how the biosynthesis is regulated. Understanding its biosynthesis and regulation will help to understand the biological function of this modification and the tumor suppression mechanism of Dph1 and Dph4, and may ultimately lead to the development of new ways to treat or prevent cancer. PUBLIC HEALTH RELEVANCE: Diphthamide is a unique post-translational modification that occurs in all eukaryotes. The biosynthesis of diphthamide requires multiple proteins, and mutations in several of them have been connected to cancer. This proposal aims to study the mechanism of diphthamide biosynthesis and the function of each protein required for the biosynthesis. Understanding the biosynthesis will provide important insight on the function of diphthamide, the regulation of diphthamide biosynthesis, and the mechanism of tumor formation in the absence of diphthamide biosynthesis, possibly leading to new ways to treat or prevent cancer.

描述(申请人提供)：敌草胺是一种翻译后修饰的组氨酸残基，存在于所有真核细胞的翻译延伸因子2(EEF-2)中。这一单一的修饰集中了许多有趣的化学和生物。这种修饰包括一个独特的C-C键形成反应，它连接了咪唑环的C2和S-腺苷蛋氨酸衍生的3-氨基-3-羧基丙基。该C-C键形成反应之后是三甲基化步骤和酰胺化步骤，以得到最终的联苯胺改性。在某些细菌感染期间，这种修饰的残基(但不是未修饰的组氨酸残基)被细菌毒素特异性识别，包括白喉毒素和假单胞菌外毒素A。这些毒素催化二磷酸腺苷核糖(ADP-核糖)从烟酰胺腺嘌呤二核苷酸(NAD)转移到咪唑环上的一个氮原子。细菌毒素的ADP核糖化作用抑制EEF-2的功能，从而抑制蛋白质的合成，导致细胞死亡。基于遗传研究，生物合成的第一步，即C-C键的形成，需要真核生物中的四种蛋白质，Dph1，Dph2，Dph3和Dph4。然而，这四种蛋白质是如何催化反应的，以及为什么有这么多蛋白质参与一个反应，目前还不清楚。Dph1和Dph4基因的杂合缺失与几种人类癌症有关，特别是卵巢癌和乳腺癌，这引发了关于敌草胺的生物学功能以及Dph1和Dph4的肿瘤抑制机制的有趣问题。在这项建议中，我们旨在研究Dph1、Dph2、Dph3和Dph4如何催化双苯二甲胺生物合成的第一步。正如我们的初步结果所预测的那样，理解生物合成将揭示一些非常有趣的酶学，并为生物合成的调控提供见解。了解其生物合成和调控将有助于了解该修饰的生物学功能以及Dph1和Dph4的抑瘤机制，并最终可能导致开发新的治疗或预防癌症的方法。公共卫生相关性：敌草胺是一种在所有真核生物中都存在的独特的翻译后修饰。敌草胺的生物合成需要多种蛋白质，其中几种蛋白质的突变与癌症有关。这项建议旨在研究敌草胺生物合成的机制和生物合成所需的每种蛋白质的功能。了解敌草胺的生物合成将为深入了解敌草胺的功能、生物合成的调控以及在没有敌草胺生物合成的情况下肿瘤的形成机制提供重要的见解，可能会为治疗或预防癌症提供新的途径。