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效率。在细菌中，在前导肽的翻译期间新生多肽介导的核糖体停滞用于诱导下游基因的表达，例如，Erm前导肽调节赋予对红霉素和其他临床上重要的大环内酯类抗生素的抗性的下游pcr基因的活性。当大环内酯类抗生素在核糖体的出口通道中结合并与新生前导肽合作以损害核糖体功能时，发生诱导BMP2基因表达所需的程序性核糖体停滞。药物依赖性停滞核糖体复合物的任何结构的缺乏限制了我们对这些调控过程的机械理解。我们最近开发了一种程序，以产生药物依赖性停滞的核糖体复合物，适用于结构审讯。该建议的目的是确定药物依赖性停滞的核糖体复合物的不同阵列的高分辨率结构。这样的结构将提供急需的结构洞察力，以阐明药物和新生肽如何协同抑制核糖体的肽键形成活性，从而诱导翻译停滞。随着病原菌多药耐药问题的日益严重，研究本文所述的药物依赖性耐药基因诱导系统将对未来新型抗菌药物的开发具有重要意义。此外，这些研究也将建立一个概念框架，了解其他辅因子和肽依赖性核糖体失速系统。

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.