Chemical Cross-linking of Protein to RNA Within Intact Ribosomal Subunits from Escherichia Coli

Overview

Journal Mol Gen Genet

Specialties Genetics
Molecular Biology

Date 1978 Apr 6

PMID 349353

Citations 13

Authors

E Ulmer

M Meinke

A Ross

G Fink

R Brimacombe

Affiliations

Soon will be listed here.

Abstract

Bifunctional reagents, namely bis-(2-chloroethyl)-amine ("nitrogen mustard") and activated esters of 3-(2-bromo-3-oxobutane-1-sulphonyl)-propionic acid ("bromo-ketone reagent") are used to cross-linked protein to RNA within intact ribosomal subunits. The cross-linked proteins are analysed on two different two-dimensional gel electrophoresis sytems, and the existence of a stable cross-linkage is demonstrated by isolating cross-linked protein-oligonucleotide complexes from subunits containing 32P-labelled RNA. Proteins S3, S4, S5, S9/S11 and S13 from the 30S subunit, and proteins L1 and L2 from the 50S subunit were cross-linked to RNA by the nitrogen mustard, together with a number of other so far unresolved proteins. Correspondingly S3, S4, S7, S9/S11 and L12 were cross-linked by the bromoketone reagent, although in lower yield. The reagents should prove useful topographical studies on ribosomal subunits, and arguments are presented favouring the use of non-cleavable and relatively non-specific RNA-protein cross-linking reagents for such studies.

Citing Articles

Chemical RNA Cross-Linking: Mechanisms, Computational Analysis, and Biological Applications.

Velema W, Lu Z JACS Au. 2023; 3(2):316-332.

PMID: 36873678 PMC: 9975857. DOI: 10.1021/jacsau.2c00625.

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.

Stiege W, Zwieb C, Brimacombe R Nucleic Acids Res. 1982; 10(22):7211-29.

PMID: 6818528 PMC: 326999. DOI: 10.1093/nar/10.22.7211.

Protein-RNA crosslinking in Escherichia coli 30S ribosomal subunits. Identification of a 16S rRNA fragment crosslinked to protein S12 by the use of the chemical crosslinking reagent 1-ethyl-3-dimethyl-aminopropylcarbodiimide.

Chiaruttini C, Hayes D, Ehresmann B Nucleic Acids Res. 1982; 10(23):7657-76.

PMID: 6760129 PMC: 327037. DOI: 10.1093/nar/10.23.7657.

Evidence for supercoiling in the DNA of bacteriophage heads.

Parish J Arch Microbiol. 1980; 126(3):277-83.

PMID: 6447487 DOI: 10.1007/BF00409932.

The use of 2-iminothiolane as an RNA-protein cross-linking agent in Escherichia coli ribosomes, and the localisation on 23S RNA of sites cross-linked to proteins L4, L6, L21, L23, L27 and L29.

Wower I, Wower J, Meinke M, Brimacombe R Nucleic Acids Res. 1981; 9(17):4285-302.

PMID: 6170935 PMC: 327435. DOI: 10.1093/nar/9.17.4285.

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