title: Photo-affinity labeling of tRNA binding sites in macromolecules. I. Linking of the phenacyl-p-azide of 4-thiouridine in (Escherichia coli) valyl-tRNA to 16S RNA at the ribosomal P site.
abstract: The phenacyl-p-azide of 4-thiouridine in (E. coli) tRNA(1) (Val) was prepared for use as a photo-affinity probe of tRNA binding sites on ribosomes. The derivatized tRNA was 90-100% as active as control tRNA for aminoacylation, nonenzymatic binding to the ribosomal P site, elongation factor Tu(EFTu)-dependent binding to the A site, EFTu-GTP-aa-tRNA ternary complex formation, and transfer of valine into polypeptide. Irradiation of p-azidophenacyl-[(3)H]valyl-tRNA bound noncovalently to the ribosomal P site resulted in covalent attachment of 15-20% of the noncovalently bound tRNA to the ribosomes. The linking occurred exclusively to the 16S RNA of the 30S ribosomal subunit, thus suggesting that the region of the ribosome within 9 A of the 4-thiouridine of tRNA, when it is bound in the P site, is solely 16S RNA.
mesh_terms: D001665:Binding Sites; D002499:Centrifugation, Density Gradient; D004926:Escherichia coli / Q000378:metabolism* / Q000648:ultrastructure; D010445:Peptide Elongation Factors; D010777:Photochemistry; D011485:Protein Binding; D011506:Proteins / Q000302:isolation & purification; D012329:RNA, Bacterial / Q000378:metabolism*; D012335:RNA, Ribosomal / Q000302:isolation & purification / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism*; D012270:Ribosomes / Q000032:analysis / Q000378:metabolism*; D013891:Thiouridine / Q000378:metabolism*; D014316:Tritium; D014633:Valine
chemical_list: D010445:Peptide Elongation Factors; D011506:Proteins; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D014316:Tritium; D013891:Thiouridine; D012343:RNA, Transfer; D014633:Valine