Does the channel for nascent peptide exist inside the ribosome? Immune electron microscopy study.

Abstract MS2 phage RNA-directed synthesis of an N-terminal polypeptide of the phage coat protein on Escherichia coli 70 S ribosomes was initiated in a cell-free system with the N -dinitrophenyl derivative of methionyl-tRNA Met F and performed in the absence of tyrosine, lysine, cysteine and methionine. As a result, the translating ribosomes carried peptides up to 42 amino acid residues in length with the dinitrophenyl hapten at the N-ends. Using the immune electron microscopy technique the positions of the nascent peptide N-ends on the 70 S ribosomes have been visualized. It has been found that (i) the N-ends of nascent peptides of these lengths are accessible to antibodies, (ii) the exit site of a nascent peptide is the pocket between the base of the central protuberance and the L1 ridge on the 50 S subunit, i.e. presumably its peptidyl transferase center, and (iii) the further pathway of a nascent peptide seems to proceed along the groove on the external surface of the 50 S subunit.