Molecular basis for drug- and peptide-dependent translational arrest

在蛋白质合成过程中，新生的多肽链可以调节翻译效率。在细菌中，在前导肽翻译过程中，新生多肽介导的核糖体停滞被用来诱导下游基因的表达，例如，Erm前导肽调节下游erm基因的活性，使其对红霉素和其他临床上重要的大环内酯类抗生素产生耐药性。诱导erm基因表达所需的程序性核糖体停滞发生在大环内酯类抗生素结合在核糖体的出口隧道中，并与新生的先导肽合作损害核糖体功能时。缺乏药物依赖的停滞核糖体复合体的任何结构限制了我们对这些调控过程的机械性理解。我们最近开发了一种方法来产生药物依赖的停滞的核糖体复合体，适用于结构询问。这项提议的目的是确定一系列药物依赖的停滞核糖体复合体的高分辨率结构。这些结构将提供急需的结构洞察力，以阐明药物和新生多肽如何合作抑制核糖体的多肽键形成活性，从而诱导翻译停滞。随着病原菌多重耐药问题的日益严重，本文所述的药物依赖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因子和多肽依赖的核糖体停滞系统建立一个概念性框架。

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.