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Molecular mechanism of protein synthesis

蛋白质合成的分子机制

基本信息

批准号：
07044183
负责人：
WATANABE Kimitsuna
金额：
$ 3.33万
依托单位：
University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for international Scientific Research
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

项目摘要

We found that a high concentration (40-60%) of pyridine, an aromatic tertiary amine catalyst, is able to promote the translation on ribosomes without soluble protein factors of chemical energy sources. This novel translation system was called the pyridine system, which could produce oligophyenylalanine with chain lengths of up to decamer and polylysine with chain lengths of around 40mer, depending on the corresponding templates, poly (U) and poly (A), respectively. In poly (UC) -dependent oligo (serine-leucine) synthesis, oligopeptides with a serine and leucine alternate sequence were the main products. The template dependency is most prominent at 60% pyridine, but not observed at 40% or 50% pyridine. The optimal temperature range of the reaction was rather wide between 30 and 60ﾟC,which may reflect no requirement for temperature-sensitive soluble protein factors for the reaction. The ranges of the optimal concentrations of K^+ (200-500 mM) and Mg^<2+> (1-50 mM) in the pyridine system … More are also rather wide, quite different from those in the usual aqueous translation system.Current investigations have suggested that rRNAs play a central role in the ribosomal functions, although the conclusive evidence has not been provided ; some papers have reported that the eubacterial peptidyltransferation, the peptide-bond formation on the ribosomal large subunits, scarcely requires their proteins. Here we demonstrate that E.coli 23S rRNAs either extracted from ribosomes or synthesized in vitro are able to promote the peptide-bond formation (phenylalanylphenylalanine synthesis from phenylalanyl-tRNA^<Phe>) without template in the presence of 40% pyridine. The reaction required 23S rRNAs or the transcripts in the folded state, and was inhibited with chloramphenicol but not with cycloheximide as well as by site-directed digestion with RNase H of 23S rRNA hybridized with some complementary oligodeoxyribonucleotides. These results strongly suggest that 23S rRNA is directly involved in the peptide bond formation. Less

我们发现，高浓度（40-60%）的吡啶，一种芳香叔胺催化剂，能够促进核糖体上的翻译，而不需要可溶性蛋白质因子的化学能源。这种新的翻译系统被称为吡啶系统，它可以产生链长高达十聚体的寡苯丙氨酸和链长约40聚体的聚赖氨酸，这取决于相应的模板，聚（U）和聚（A），分别。在多聚（UC）依赖性寡（丝氨酸-亮氨酸）合成中，具有丝氨酸和亮氨酸交替序列的寡肽是主要产物。模板依赖性在60%吡啶时最显著，但在40%或50%吡啶时未观察到。反应的最适温度范围在30 ~ 60 ℃之间，这可能反映了反应不需要温度敏感的可溶性蛋白因子。吡啶体系中K^+（200-500 mM）和Mg^<2+>（1-50 mM）的最佳浓度范围 ...更多信息目前的研究表明，rRNA在核糖体功能中起着核心作用，尽管尚未提供确凿的证据;一些文献报道，真细菌的肽基转移，即核糖体大亚基上的肽键形成，几乎不需要它们的蛋白质。在这里，我们证明了从核糖体中提取的或体外合成的大肠杆菌23 S rRNA能够在40%吡啶存在下在没有模板的情况下促进肽键形成（从苯丙氨酰-tRNA ^合成苯丙氨酰苯丙氨酸<Phe>）。该反应需要23 S rRNA或在折叠状态的转录本，并抑制氯霉素，但不与放线菌酮，以及通过与一些互补的寡脱氧核苷酸杂交的23 S rRNA的RNA酶H定点消化。这些结果有力地表明，23 S rRNA直接参与了肽键的形成。少