Crosslinking of chemically reactive A-U-G analogs to ribosomal proteins within initiation complexes. |
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Authors: | O. Pongs H.U. Petersen M. Grunberg-Manago E. Lanka R. Bald G. Stöffler |
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Affiliation: | Ruhr-Universität Bochum, Lehrstuhl für Biochemie 4630 Bochum 1, West Germany;Institut de Biologie Physicochemique, Paris, France;Max-Planck-Institut für Molekulare Genetik, Berlin, West Germany |
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Abstract: | 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. |
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Keywords: | To whom correspondence should be addressed. |
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