PROBING RIBOSOMAL FUNCTION

本研究的主要目的是阐明 核糖体特别是暴露的单链 核糖体RNA的区域将被分析，以确定它们如何影响 核糖体的功能。通过这种方式，tRNA、MRNA和 可以确定翻译因子绑定。此外， 核糖体在翻译中的功能可以是RNA的结构。 观察特别令人感兴趣的是抗生素结合的位点， 由于核糖体功能或结构在这些细胞中受到显著影响， 例实验的设计是化学合成短 与核糖体RNA的特定区域互补的DNA寡聚体， 将这些DNA探针与核糖体上的靶位点杂交， 对tRNA结合、mRNA结合、翻译因子 结合或抗生素结合。通过以下方法测定相对结合率 监测放射性标记探针的残余结合活性 或配体。研究中心特异性 将使用RNaseH测定结合。利用这些结果， 开始了解mRNA、tRNA、翻译因子 而核糖体则负责合成蛋白质。

The overriding purpose of this research is to elucidate the function of the ribosome. Specifically the function of exposed, single-stranded regions of ribosomal RNA will be assayed to determine how they affect the function of the ribosome. In this way the sites where TRNA, MPNA and translation factors bind can be determined. In addition, changes in the structure of the RNA as the ribosome functions in translation can be observed. Of particular interest are the sites where antibiotics bind, since ribosomal function or structure is markedly affected in these cases. The design of the experiments is to chemically synthesize short DNA oligomers complementary to specific regions of ribosomal RNA, hybridize these DNA probes to target sites on the ribosome, and assay the resulting affects on tRNA binding, mRNA binding, translation factor binding or antibiotic binding. Relative binding will be assayed by monitoring the residual binding activity of radioactively-labeled probes or ligands using nitrocellulose filter assays. Specificity of site binding will be determined using RNase H. Using these results it should be possible to begin to understand how mRNA, tRNA, translation factors and the ribosome go about the business of synthesizing proteins.