Douthwaite S, Christensen A, Garrett R A
Biochemistry. 1982 May 11;21(10):2313-20. doi: 10.1021/bi00539a007.
The binding sites of ribosomal proteins L18 and L25 on 5S RNA from Escherichia coli were probed with ribonucleases A, T1, and T2 and a double helix specific cobra venom endonuclease. The results for the protein-RNA complexes, which were compared with those for the free RNA [Douthwaite, S., & Garrett, R. A. (1981) Biochemistry 20, 7301--7307], reveal an extensive interaction site for protein L18 and a more localized one for L25. Generally comparable results, with a few important differences, were obtained in a study of the binding sites of the two E. coli proteins on Bacillus stearothermophilus 5S RNA. Several protein-induced changes in the RNA structures were identified; some are possibly allosteric in nature. The two prokaryotic 5S RNAs were also incubated with total 50S subunit proteins from E. coli and B. stearothermophilus ribosomes. Homologous and heterologous reconstitution experiments were performed for both RNAs. The effects of the bound proteins on the ribonuclease digestion of the RNAs could generally be correlated with the results obtained with the E. coli proteins L18 and L25, although there was evidence for an additional protein-induced conformational change in the B. stearothermophilus 5S RNA, which may have been due to a third ribosomal protein L5.
利用核糖核酸酶A、T1、T2以及一种双链螺旋特异性眼镜蛇毒内切酶，对核糖体蛋白L18和L25在大肠杆菌5S RNA上的结合位点进行了探测。将蛋白质-RNA复合物的结果与游离RNA的结果[杜思韦特，S.，&加勒特，R. A.（1981年）《生物化学》20，7301 - 7307]进行比较，结果显示蛋白质L18存在一个广泛的相互作用位点，而L25的相互作用位点则更为局限。在一项关于这两种大肠杆菌蛋白质在嗜热栖热放线菌5S RNA上结合位点的研究中，得到了大致可比的结果，但也存在一些重要差异。确定了RNA结构中几种由蛋白质诱导的变化；其中一些可能具有变构性质。还将这两种原核生物的5S RNA与来自大肠杆菌和嗜热栖热放线菌核糖体的完整50S亚基蛋白质一起孵育。对这两种RNA都进行了同源和异源重组实验。结合的蛋白质对RNA核糖核酸酶消化的影响通常与用大肠杆菌蛋白质L18和L25得到的结果相关，尽管有证据表明嗜热栖热放线菌5S RNA中存在一种额外的由蛋白质诱导的构象变化，这可能是由于第三种核糖体蛋白L5所致。
