Molecular basis for drug- and peptide-dependent translational arrest

在蛋白质合成过程中，新生多肽链可以调节翻译的效率。在细菌中，在先导肽翻译过程中，新生多肽介导的核糖体停顿被用来诱导下游基因的表达，例如，Erm先导肽调节下游Erm基因的活性，从而赋予对红霉素和其他临床重要的大环内酯类抗生素的抗性。当大环内酯类抗生素结合在核糖体的出口通道中并与新生先导肽合作损害核糖体功能时，诱导erm基因表达所需的程序性核糖体延迟发生。缺乏任何药物依赖的停滞核糖体复合物的结构限制了我们对这些调节过程的机制理解。我们最近开发了一种程序来产生药物依赖的停滞核糖体复合物，适用于结构审讯。本提案的目的是确定高分辨率结构的各种药物依赖的停滞核糖体复合物。这种结构将为阐明药物和新生肽如何协同抑制核糖体的肽键形成活性从而诱导翻译阻滞提供急需的结构见解。随着病原菌多药耐药问题的日益严重，本文所描述的药物依赖性耐药基因诱导系统的研究将对未来新型抗菌药物的开发具有重要意义。此外，这些研究还将为理解其他辅助因子和肽依赖的核糖体延迟系统建立一个概念框架。

During protein synthesis, nascent polypeptide chains can modulate the efficiency of translation. In bacteria, nascent polypeptide-mediated ribosome stalling during translation of leader peptides is used to induce expression of downstream genes, for example, Erm leader peptides modulate activity of the downstream erm genes that confer resistance to erythromycin and other clinically-important macrolide antibiotics. The programmed ribosome stalling required for induction of erm gene expression occurs when the macrolide antibiotic binds in the exit tunnel of the ribosome and cooperates with the nascent leader peptide to impair ribosomal function. The absence of any structures of drug-dependent stalled ribosome complexes has limited our mechanistic understanding of these regulatory processes. We have recently developed a procedure to generate drug-dependent stalled ribosome complexes that are suitable for structural interrogation. The aim of this proposal is to determine high resolution structures of a diverse array of drug-dependent stalled ribosome complexes. Such structures will provide much needed structural insight to elucidate how the drug and the nascent peptide cooperate to inhibit the peptide bond formation activity of the ribosome and thereby induce translation arrest. With the ever-increasing problem of multi-drug resistance in pathogenic bacteria, insight gained from investigating the drug-dependent resistance gene induction systems described here will be important for the future development of novel antimicrobial agents. Moreover, such studies will also establish a conceptual framework for understanding other cofactor- and peptide-dependent ribosome stalling systems.