Cooperative interactions of RNA and thiostrepton antibiotic with two domains of ribosomal protein L11.
Ribosomal protein L11 interacts with a 58-nucleotide domain of large subunit ribosomal RNA; both the protein and its RNA target have been highly conserved. The antibiotic thiostrepton recognizes the same RNA domain, and binds to the ribosome cooperatively with L11. Experiments presented here show that RNA recognition and thiostrepton cooperativity can be attributed to C- and N-terminal domains of L11, respectively. Under trypsin digestion conditions that degrade Bacillus stearothermophilus L11 to small fragments, the target RNA protects the C-terminal 77 residues from digestion, and thiostrepton and RNA in combination protect the entire protein. A 76-residue C-terminal fragment of L11 was overexpressed and shown to fold into a stable structure binding ribosomal RNA with essentially the same properties as full-length L11. An L11.thiostrepton.RNA complex was 100-200-fold more stable than expected on the basis of L11-RNA and thiostrepton-RNA binding affinities; similar measurements with the C-terminal fragment detected no cooperativity with thiostrepton. L11 function is thus more complex than simple interaction with ribosomal RNA; we suggest that thiostrepton mimics some ribosomal component or factor that normally interacts with the L11 N-terminal domain.
Xing
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Department of Chemistry, Johns Hopkins University, Baltimore, Maryland 21218, USA.
Draper
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D000900:Anti-Bacterial Agents / Q000378:metabolism*; D001411:Geobacillus stearothermophilus; D008968:Molecular Conformation; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D010446:Peptide Fragments / Q000378:metabolism; D012338:RNA, Ribosomal, 23S / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism; D012269:Ribosomal Proteins / Q000378:metabolism*; D013883:Thiostrepton / Q000378:metabolism*; D014357:Trypsin
D000900:Anti-Bacterial Agents; D010446:Peptide Fragments; D012338:RNA, Ribosomal, 23S; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins; C027624:ribosomal protein L11; D014357:Trypsin; D013883:Thiostrepton