Mapping the path of the nascent peptide chain through the 23S in the 50S ribosomal subunit

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Abstract
Peptides of different lengths encoded by suitable mRNA fragments were biosyntheslzed in situ on Escherichia coll rlbosomes. The peptides carried a diazirine derivative bound to their N-terminal methlonlne residue, which was photoactivated whilst the peptides were still attached to the ribosome. Subsequently, the sites of photo-cross-linking to 23S RNA were analyzed by our standard procedures. The N-termini of peptides of increasing length became progressively cross-linked to nucleotlde 750 (peptides of 6, 9 or 13–15 amino acids), to nucleotide 1614 and concomltantly to a second site between nucleotides 1305 and 1350 (a peptide of 25–26 amino acids), and to nucleotide 91 (a peptide of 29–33 amino acids). reviously we had shown that peptides of 1 or 2 amino acids were cross-linked to nucleotides 2062,2506 and 2585 within the peptidyl transferase ring, whereas triand tetrapeptides were additionally cross-linked to nucleotides 2609 and 1781. Taken together, the data demonstrate that the path of the nascent peptide chain moves from the peptidyl transferase ring in domain V of the 23S RNA to domain IV, then to domain II, then to domain III, and finally to domain I. These cross-linking results are correlated with other types of topographical data relating to the 50S subunit