Peptide inhibitors of peptidyltransferase alter the conformation of domains IV and V of large subunit rRNA: a model for nascent peptide control of translation.
Peptides of 5 and 8 residues encoded by the leaders of attenuation regulated chloramphenicol-resistance genes inhibit the peptidyltransferase of microorganisms from the three kingdoms. Therefore, the ribosomal target for the peptides is likely to be a conserved structure and/or sequence. The inhibitor peptides "footprint" to nucleotides of domain V in large subunit rRNA when peptide-ribosome complexes are probed with dimethyl sulfate. Accordingly, rRNA was examined as a candidate for the site of peptide binding. Inhibitor peptides MVKTD and MSTSKNAD were mixed with rRNA phenol-extracted from Escherichia coli ribosomes. The conformation of the RNA was then probed by limited digestion with nucleases that cleave at single-stranded (T1 endonuclease) and double-stranded (V1 endonuclease) sites. Both peptides selectively altered the susceptibility of domains IV and V of 23S rRNA to digestion by T1 endonuclease. Peptide effects on cleavage by V1 nuclease were observed only in domain V. The T1 nuclease susceptibility of domain V of in vitro-transcribed 23S rRNA was also altered by the peptides, demonstrating that peptide binding to the rRNA is independent of ribosomal protein. We propose the peptides MVKTD and MSTSKNAD perturb peptidyltransferase center catalytic activities by altering the conformation of domains IV and V of 23S rRNA. These findings provide a general mechanism through which nascent peptides may cis-regulate the catalytic activities of translating ribosomes.
Harrod
R
R
Department of Biological Sciences, University of Maryland, Catonsville 21228, USA.
Lovett
P S
PS
D000595:Amino Acid Sequence; D001483:Base Sequence; D004720:Endonucleases / Q000378:metabolism; D008957:Models, Genetic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009842:Oligopeptides / Q000378:metabolism / Q000494:pharmacology*; D010442:Peptide Chain Initiation, Translational; D010458:Peptidyl Transferases / Q000037:antagonists & inhibitors*; D011485:Protein Binding; D014176:Protein Biosynthesis; D012338:RNA, Ribosomal, 23S / Q000187:drug effects* / Q000378:metabolism
D009842:Oligopeptides; D012338:RNA, Ribosomal, 23S; D010458:Peptidyl Transferases; D004720:Endonucleases