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通过30S核糖体亚基内的原位RNA内交联研究大肠杆菌16S RNA的三级折叠。

Investigation of the tertiary folding of Escherichia coli 16S RNA by in situ intra-RNA cross-linking within 30S ribosomal subunits.

作者信息

Atmadja J, Brimacombe R, Blöcker H, Frank R

出版信息

Nucleic Acids Res. 1985 Oct 11;13(19):6919-36. doi: 10.1093/nar/13.19.6919.

DOI:10.1093/nar/13.19.6919
PMID:2997731
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC322013/
Abstract

Intra-RNA cross-links were introduced into E. coli 30S ribosomal subunits by mild ultraviolet irradiation. The subunits were partially digested with cobra venom nuclease, followed in some cases by a second partial digestion with ribonuclease H in the presence of the hexanucleotide d-(CTTCCC). The cross-linked RNA complexes were separated by two-dimensional gel electrophoresis and the sites of cross-linking analysed by our published procedures. Tertiary structural cross-links in the 16S RNA were identified between positions 31 and 48, between oligonucleotides 1090-1094 and 1161-1164, and between oligonucleotides 1125-1127 and 1280-1281. The first of these imposes a rigid constraint on the relative orientations of helices 3 and 4 of the 16S secondary structure. A further tertiary cross-link (which could not be precisely localised) was found between regions 1-72 and 1020-1095, and secondary structural cross-links were identified between positions 497 and 545-548, and positions 1238-1240 and 1298.

摘要

通过温和紫外线照射将RNA内交联引入大肠杆菌30S核糖体亚基。用眼镜蛇毒核酸酶对亚基进行部分消化，在某些情况下，随后在六核苷酸d-(CTTCCC)存在下用核糖核酸酶H进行第二次部分消化。通过二维凝胶电泳分离交联的RNA复合物，并按照我们已发表的方法分析交联位点。在16S RNA中，在第31和48位之间、寡核苷酸1090 - 1094和1161 - 1164之间以及寡核苷酸1125 - 1127和1280 - 1281之间鉴定出三级结构交联。其中第一个对16S二级结构的螺旋3和4的相对取向施加了严格限制。在区域1 - 72和1020 - 1095之间发现了另一个三级交联（无法精确定位），并在第497和545 - 548位之间以及第1238 - 1240和1298位之间鉴定出二级结构交联。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/158e14a02677/nar00313-0140-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/629052a78d63/nar00313-0137-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/3629e963613f/nar00313-0138-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/158e14a02677/nar00313-0140-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/629052a78d63/nar00313-0137-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/3629e963613f/nar00313-0138-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f31d/322013/158e14a02677/nar00313-0140-a.jpg

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Precise localisation of three intra-RNA cross-links in 23S RNA and one in 5S RNA, induced by treatment of Escherichia coli 50S ribosomal subunits with bis-(2-chloroethyl)-methylamine.用双（2-氯乙基）甲胺处理大肠杆菌50S核糖体亚基后，在23S RNA中诱导产生的三个RNA内交联以及在5S RNA中诱导产生的一个交联的精确定位。
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