Modification, inhibition and function of elongation factor EF-P

伸长因子EF-P的修饰、抑制及其功能

• 批准号: 265881756
• 负责人: Professorin Dr. Kirsten Jung
• 金额: --
• 依托单位: Abteilung Mikrobiologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2018-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/265881756?language=en
• 关键词: Modification; inhibition; function; elongation; factor

The fundamental process of protein synthesis is catalyzed on ribosomes. We have recently demonstrated that bacterial ribosomes become arrested when translating proteins contain consecutive polyproline stretches, and that this arrest is alleviated by the translation elongation factor EF-P. Furthermore, we could show that the post-translational ß-lysinylation of lysine34 of Escherichia coli EF-P by the enzymes YjeA and YjeK and the rhamnosylation of arginine32 of Shewanella EF-P by EarP, respectively, are critical for the rescue activity of EF-P. Many virulence factors contain polyproline stretches, explaining why modified EF-P is critical for bacterial pathogenicity. While EF-P is conserved, the modification enzymes YjeA and YjeK or EarP are absent in many bacterial phyla, even though these bacteria encode a high number of polyproline-containing proteins, thus suggesting the existence of novel EF-P modification pathways. Here we propose to identify and characterize these novel modification pathways, with a particular focus on phyla containing clinically important pathogenic bacteria, such as Firmicutes (e.g., Staphylococcus and Enterococcus) and Actinobacteria (e.g., Mycobacterium). In addition, we propose to apply an in vivo screening assay to identify low molecular weight EF-P inhibitors for the development of novel antimicrobial agents. Lastly, we aim to resolve structures of modified EF-P bound to polyproline-stalled ribosomes in order to elucidate the mechanism by which polyproline stretches stall ribosomes as well as how EF-P alleviates the translational arrest. Collectively, we believe our proposal will not only provide new functional and mechanistic insights into the fundamental process of translation, but also extend the spectrum of post-translational modification systems in general.

蛋白质合成的基本过程是在核糖体的催化下进行的。我们最近证明，当翻译蛋白质包含连续的聚脯氨酸延伸时，细菌核糖体会被停滞，并且这种停滞可以通过翻译延伸因子 EF-P 得到缓解。此外，我们还可以证明，酶 YjeA 和 YjeK 对大肠杆菌 EF-P 的赖氨酸 34 进行翻译后 β-赖氨酰化，以及 EarP 对希瓦氏菌 EF-P 的精氨酸 32 进行翻译后鼠李糖基化，这对于 EF-P 的救援活性至关重要。许多毒力因子含有聚脯氨酸片段，这解释了为什么修饰的 EF-P 对于细菌致病性至关重要。虽然 EF-P 是保守的，但许多细菌门中不存在修饰酶 YjeA 和 YjeK 或 EarP，即使这些细菌编码大量含聚脯氨酸的蛋白质，因此表明存在新的 EF-P 修饰途径。在这里，我们建议识别和表征这些新的修饰途径，特别关注含有临床重要病原菌的门，例如厚壁菌门（例如葡萄球菌和肠球菌）和放线菌（例如分枝杆菌）。此外，我们建议应用体内筛选试验来鉴定低分子量 EF-P 抑制剂，用于开发新型抗菌药物。最后，我们的目标是解析与聚脯氨酸失速核糖体结合的修饰 EF-P 的结构，以阐明聚脯氨酸拉伸失速核糖体的机制以及 EF-P 如何减轻翻译停滞。总的来说，我们相信我们的建议不仅将为翻译的基本过程提供新的功能和机制见解，而且还将扩展翻译后修饰系统的范围。