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Real-time evidence for EF-G-induced dynamics of helix 44 in 16S rRNA

机译：EF-G诱导16S rRNA中44螺旋动力学的实时证据

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摘要

The penultimate stem-loop of 16S ribosomal RNA (rRNA), helix 44, plays a central role in ribosome function. Using time-resolved dimethyl sulfate (DMS) probing, we have analyzed time-dependent modifications that occur at specific bases in this helix near the decoding region, resulting from the binding of elongation factor G (EF-G) in various forms. When EF-G-GTP is bound to 70S ribosomes, bases A1492 and A1493 are immediately protected, while other bases in the region show either no change or enhanced modification. When apo-EF-G is bound to 70S ribosomes and GTP is added, substantial transient time-dependent enhancement occurs at bases A1492 and A1493, with somewhat less enhancement occurring at base A1483, all in the first 45 ms. When mRNA and deacylated tRNAs are bound to the 70S ribosome and EF-G-GTP is added, bases A1492 and A1493 again show substantial and continued enhancement, while bases A1408, A1413, and A1418 all show time-dependent protection. These results provide primary, real-time evidence that EF-G induces direct or indirect structural changes in this region as EF-G is bound and as GTP is hydrolyzed.
机译：16S核糖体RNA（rRNA）的倒数第二个茎环，螺旋线44在核糖体功能中起着核心作用。使用时间分辨的硫酸二甲酯（DMS）探测，我们分析了由于多种形式的延伸因子G（EF-G）结合而在此螺旋附近解码区特定碱基处发生的时间依赖性修饰。当EF-G-GTP与70S核糖体结合时，碱基A1492和A1493立即受到保护，而该区域中的其他碱基则无变化或修饰增强。当apo-EF-G与70S核糖体结合并添加GTP时，在第一个45毫秒内，碱基A1492和A1493发生了明显的瞬时时间依赖性增强，而碱基A1483出现了较少的增强。当mRNA和脱酰tRNA结合到70S核糖体并添加EF-G-GTP时，碱基A1492和A1493再次显示出实质性且持续的增强，而碱基A1408，A1413和A1418均显示出时间依赖性。这些结果提供了主要的实时证据，表明当EF-G结合且GTP水解时，EF-G会在该区域诱导直接或间接的结构变化。

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