Flexibility of the nascent polypeptide chain within the ribosome--contacts from the peptide N-terminus to a specific region of the 30S subunit.
The ribosomal environment of the N-terminus of the nascent polypeptide chain has been investigated using peptides of different lengths, synthesized in situ on Escherichia coli ribosomes; the peptides each carry a photoreactive diazirine moiety at their N-terminus, so as to generate cross-links to neighbouring ribosomal components. Our previous studies [Choi, K. M. & Brimacombe, R. (1998) Nucleic Acids Res. 26, 887-895] with three independent families of peptides, derived from the E. coli ompA protein gene, the tetracycline-resistance gene and the bacteriophage T4 gene 60, identified a series of sites within the 23S rRNA to which the peptides became cross-linked. The distribution of these cross-links indicated that the nascent peptide is very flexible within the 50S subunit. Here, we demonstrate that the N-termini of the ompA and gene-60 peptides can, in addition, even become concomitantly cross-linked to the 30S subunit. The cross-linking is predominantly to 30S ribosomal proteins S1, S2, S4 and (to a lesser extent) S3, which form a cluster near to the decoding region. This result is discussed in terms of the flexibility of the nascent peptide during the co-translational folding process, and in terms of the 'ribosomal bypass' phenomenon which is known to occur during translation of the gene 60 mRNA.
Choi
K M
KM
Max-Planck-Institut für Molekulare Genetik, Berlin, Germany.
Atkins
J F
JF
Gesteland
R F
RF
Brimacombe
R
R
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis / Q000737:chemistry; D017122:Bacteriophage T4 / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000737:chemistry*; D012338:RNA, Ribosomal, 23S / Q000737:chemistry / Q000378:metabolism*; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D013753:Tetracycline Resistance / Q000235:genetics
D001425:Bacterial Outer Membrane Proteins; D010446:Peptide Fragments; D012338:RNA, Ribosomal, 23S; C078167:OMPA outer membrane proteins