SELECTION OF THE MESSENGER-RNA TRANSLATION INITIATION REGION BY ESCHERICHIA-COLI RIBOSOMES
SELECTION OF THE MESSENGER-RNA TRANSLATION INITIATION REGION BY ESCHERICHIA-COLI RIBOSOMES
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DOI：
10.1073/pnas.85.17.6427
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发表时间：
1988-09-01
影响因子：
11.1
通讯作者：
GUALERZI, CO
中科院分区：
文献类型：
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作者：
CALOGERO, RA;PON, CL;GUALERZI, CO
组装、克隆和高产量转录指定最小大小和编码能力的模型mRNA（具有或不具有Shine-Dalgarno（SD）序列）的两个基因。这些mRNA以及合成的多核苷酸、噬菌体MS 2 RNA和与16 S rRNA的3“末端互补的脱氧八核苷酸用于研究体外翻译起始的机制。大肠杆菌30 S核糖体亚基与所有这些核酸相互作用，尽管亲和力不同; mRNA与SD序列的亲和力（Ka. apprxeq. 2倍。107 M-1）比缺乏该序列的mRNA高一个数量级以上。起始因子对于30 S和70 S起始复合物的形成和mRNA翻译同样是必需的，无论SD序列的存在与否，但起始因子不影响SD相互作用或mRNA与核糖体的结合。SD相互作用也与30 S起始复合物的形成机制无关，并且对于体外翻译或mRNA阅读框架的选择不是必需的。有人建议，SD相互作用的功能是为了确保高浓度的起始三联体附近的核糖体肽基-tRNA结合位点，而翻译开始的选择实现动力学，在起始因子的影响下，在解码的起始tRNA。
Two genes specifying model mRNAs of minimal size and coding capacity, with or without the Shine-Dalgarno (SD) sequence, were assembled, cloned, and transcribed in high yields. These mRNAs, as well as synthetic polynucleotides, phage MS2 RNA, and a deoxyoctanucleotide complementary to the 3'' end of 16S rRNA were used to study the mechanism of translation initiation in vitro. Escherichia coli 30S ribosomal subunits interact with all these nucleic acid, albeit with different affinities; the affinity for the mRNA with the SD sequence (Ka .apprxeq. 2 .times. 107 M-1) is more than an order of magnitude higher than that for the mRNA lacking this sequence. The initiation factors are equally required, regardless of the presence of the SD sequence, for 30S and 70S initiation complex formation and for mRNA translation, but the initiation factors do not affect the SD interaction or the binding of the mRNAs to the ribosomes. The SD interaction is also mechanistically irrelevant for 30S initiation complex formation and is not essential for translation in vitro or for the selection of the mRNA reading frame. It is suggested that the function of the SD interaction is to ensure a high concentration of the initiation triplet near the ribosomal peptidyl-tRNA binding site, whereas the selection of the translational start is achieved kinetically, under the influence of the initiation factors, during decoding of the initiator tRNA.