CROSSLINKING OF CHEMICALLY REACTIVE A-U-G ANALOGS TO RIBOSOMAL-PROTEINS WITHIN INITIATION-COMPLEXES

A-U-G analogs, either reactive on their 5′ or their 3′ side, were employed in affinity labeling of the ribosomal A-U-G binding site. These experiments have been carried out such that the chemically reactive A-U-G analog became covalently bonded to ribosomal proteins only in the presence of fMet-tRNAfMet and initiation factors. Subsequent radioimmunodiffusion of A-U-G-labeled proteins identified proteins IF3, S1, S18, S21 and L11 as being in the neighborhood of the ribosomal codon binding site. A location of reactive sites of these proteins relative to the P or A site bound codon is, however, not clear. The A-U-G labeling results are quantitatively as well as qualitatively very different in the absence or presence of fMet-tRNAfMet. It is concluded, therefore, that fMet-tRNAfMet directs A-U-G into its final binding site. Streptomycin cannot release fMet-tRNAfMet from initiation complexes which contain irreversibly bound 5′- {4-(bromoacetamido)phenylphospho}-adenylyl-(3′-5′)-uridylyl-(3′-5′)-guanosine. This suggests that codon-anticodon interaction between A-U-G and fMet-tRNAfMet is still intact in the P site of the ribosome. © 1979.