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The path of mRNA through the bacterial ribosome: a site-directed crosslinking study using new photoreactive derivatives of guanosine and uridine
- Source :
- Scopus-Elsevier
- Publication Year :
- 1997
-
Abstract
- Two new photoreactive nucleotide derivatives have been applied in site-directed crosslinking studies with mRNA analogues. 6-Thioguanosine triphosphate or 5-methyleneaminouridine triphosphate was incorporated into mRNA analogues by T7 transcription; after transcription, the 5-methyleneaminouridine residues were converted to a diazirine derivative. mRNA analogues carrying either 6-thioguanosine or the diazirine derivative were bound to Escherichia coli ribosomes in the presence of tRNA(f)(Met), and photo-crosslinking was induced by irradiation at 350 nm. With 6-thioguanosine, specific crosslinks were observed from downstream positions +8 or +9 of the mRNA to nt 1196 in helix 34 of the 16S rRNA, and from position +12 to nt 530 in helix 18. With the diazirine derivative, a crosslink from position +2 (within the AUG codon) to nt 926 in helix 28 was found. Taken together with previous data obtained from downstream sites in mRNA analogues carrying 4-thiouridine residues, specific crosslinks have now been identified from downstream mRNA positions +2, +4, +6, +7, +8, +9, +11, and +12. The data confirm that the three 16S rRNA regions involved-helices 18, 28, and 34-are in the direct neighborhood of the decoding area of the 30S subunit.
- Subjects :
- RNA, Transfer, Met
Base Sequence
Transcription, Genetic
Molecular Sequence Data
Uridine Triphosphate
Thionucleotides
RNA, Bacterial
Cross-Linking Reagents
Bacteriophage T7
RNA, Ribosomal, 16S
Escherichia coli
Nucleic Acid Conformation
Guanosine Triphosphate
RNA, Messenger
Codon
Ribosomes
Research Article
Subjects
Details
- Language :
- English
- Database :
- OpenAIRE
- Journal :
- Scopus-Elsevier
- Accession number :
- edsair.pmid.dedup....452e33e06cfa1257d93e43c8e704790f