Department of Chemistry and A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119899, Russia.
J Mol Biol. 2012 Mar 9;416(5):656-67. doi: 10.1016/j.jmb.2012.01.005. Epub 2012 Jan 10.
Catalysis of peptide bond formation in the peptidyl transferase center is a major enzymatic activity of the ribosome. Mutations limiting peptidyl transferase activity are mostly lethal. However, cellular processes triggered by peptidyl transferase deficiency in the bacterial cell are largely unknown. Here we report a study of the lethal G2061C mutant of Escherichia coli 23S ribosomal RNA (rRNA). The G2061C mutation completely impaired the puromycin reaction and abolished formation of the active firefly luciferase in an in vitro translation system, while poly(U)- and short synthetic mRNA-directed peptidyl transferase reaction with aminoacylated tRNAs in vitro was seemingly unaffected. Study of the cellular proteome upon expression of the 23S rRNA gene carrying the G2061C mutation compared to cells expressing wild-type 23S rRNA gene revealed substantial differences. Most of the observed effects in the mutant were associated with reduced expression of stress response proteins and particularly proteins associated with the ppGpp-mediated stringent response.
肽键形成的催化在肽基转移酶中心是核糖体的主要酶活性。限制肽基转移酶活性的突变大多是致命的。然而，细菌细胞中肽基转移酶缺乏所引发的细胞过程在很大程度上是未知的。在这里，我们报告了对大肠杆菌 23S 核糖体 RNA(rRNA)的致命 G2061C 突变体的研究。G2061C 突变完全抑制了嘌呤霉素反应，并在体外翻译系统中消除了活性萤火虫荧光素酶的形成，而体外多聚(U)-和短合成 mRNA 指导的肽基转移酶反应与氨酰化 tRNA 似乎不受影响。与表达野生型 23S rRNA 基因的细胞相比，在表达携带 G2061C 突变的 23S rRNA 基因的细胞中研究细胞蛋白质组时，发现了明显的差异。在突变体中观察到的大多数影响与应激反应蛋白的表达减少有关，特别是与 ppGpp 介导的严格反应相关的蛋白。
