Topography of the E site on the Escherichia coli ribosome.
Topography of the E site on the Escherichia coli ribosome.
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大肠杆菌核糖体上 E 位点的地形图。
DOI：
10.1002/j.1460-2075.1993.tb05694.x
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Zimmermann,RA
中科院分区：
文献类型：
--
作者：
Wower,J;Scheffer,P;Sylvers,LA;Wintermeyer,W;Zimmermann,RA
三个光反应性tRNA探针已被用于鉴定与氨基酰基受体末端和与大肠杆菌核糖体E位点结合的tRNA的反密码子环接触的核糖体组分。其中两个探针是E. coli tRNA（Phe），其中位置73和76处的腺苷被2-叠氮基腺苷替换。第三探针通过用2-叠氮腺苷取代位置37处的尿苷而衍生自酵母tRNA（Phe）。尽管进行了修饰，但所有的光反应性tRNA种类都能够结合到E.大肠杆菌核糖体程序与聚（A），并在照射后，形成共价加合物与核糖体亚单位。在3′端或附近修饰的tRNA（Phe）探针仅标记50 S亚基中的蛋白L33。在反密码子环内含有2-叠氮腺苷的tRNA（Phe）衍生物与蛋白质S11以及包含3′末端30个核苷酸的16 S rRNA片段交联。根据免疫电镜确定的16 SrRNA的3′端和L33、S11的位置，对E位点结合的tRNA在核糖体亚基上的两端进行了定位。我们的研究结果最终表明，E网站是地形不同的P网站或A网站，它位于旁边的P网站预期的tRNA出口网站。
Three photoreactive tRNA probes have been utilized in order to identify ribosomal components that are in contact with the aminoacyl acceptor end and the anticodon loop of tRNA bound to the E site of Escherichia coli ribosomes. Two of the probes were derivatives of E. coli tRNA(Phe) in which adenosines at positions 73 and 76 were replaced by 2‐azidoadenosine. The third probe was derived from yeast tRNA(Phe) by substituting wyosine at position 37 with 2‐azidoadenosine. Despite the modifications, all of the photoreactive tRNA species were able to bind to the E site of E. coli ribosomes programmed with poly(A) and, upon irradiation, formed covalent adducts with the ribosomal subunits. The tRNA(Phe) probes modified at or near the 3′ terminus exclusively labeled protein L33 in the 50S subunit. The tRNA(Phe) derivative containing 2‐azidoadenosine within the anticodon loop became cross‐linked to protein S11 as well as to a segment of the 16S rRNA encompassing the 3′‐terminal 30 nucleotides. We have located the two extremities of the E site‐bound tRNA on the ribosomal subunits according to the positions of L33, S11 and the 3′ end of 16S rRNA defined by immune electron microscopy. Our results demonstrate conclusively that the E site is topographically distinct from either the P site or the A site, and that it is located alongside the P site as expected for the tRNA exit site.