Ribosomal proteins S7 and L1 are located close to the decoding site of E. coli ribosome ― affinity labeling studies with modified tRNAs carrying photoreactive probes attached adjacent to the 3'-end of the anticodon

• Published in: Nucleic acids research Vol. 17; no. 21; pp. 8767 - 8782
• Main Authors: PODKOWINSKI, J, GORNICKI, P
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 11.11.1989
• Subjects: Affinity Labels; Anticodon; Azides; Bacterial Proteins - genetics; Biological and medical sciences; Cross-Linking Reagents; Escherichia coli; Escherichia coli - metabolism; Fundamental and applied biological sciences. Psychology; Molecular and cellular biology; Molecular Conformation; Molecular genetics; Ribosomal Proteins - genetics; Ribosomes - metabolism; RNA, Transfer; RNA, Transfer, Amino Acid-Specific - genetics; RNA, Transfer, Met - genetics; Translation. Translation factors. Protein processing; Ligand binding; Ribosome; Radiolabelling; Escherichia coli; Crosslinking; Photoaffinity labelling; Ribosomal protein; Sucrose gradient; Chemical modification; t-RNA; Bacteria; Localization; Escherichieae; Anticodon; Enterobacteriaceae
• ISSN: 0305-1048; 1362-4962

Two photoreactive azidonitrophenyl probes have been attached to Yeast methionine elongator tRNA by chemical modification of the N6-(threoninocarbonyl)adenosine located next to the 3'-end of the anticodon. The maximum distance between the purine ring and the azido group estimated for the two probes is 16-17 and 23-24A, respectively. Binding and cross-linking of the uncharged, modified tRNAs to E. coli ribosomes have been studied with and without poly(A,U,G) as a message, under conditions directing uncharged tRNAs preferentially to the P-site. The modified tRNAs retain their binding activity and upon irradiation bind covalently to the ribosome with very high yields. Protein S7 is the major cross-linking target for both modified tRNAs, in the presence or absence of poly(A,U,G). Protein L1 and to a lesser extent proteins L33 and L27 have been found to be cross-linked with the short probe. Cross-linking to 168 rRNA reaches significant levels only in the absence of the message.