Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 288, Issue 26, Pages 19081-19089Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M113.466748
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Funding
- Swedish Research Council (M section)
- Swedish Research Council (NT section)
- Swedish Research Council (VR-SIDA)
- Carl Tryggers Stiftelse
- Wenner Gren Stiftelse
- Knut and Wallice Wallenberg Foundation
- Vinnova/DBT (India)
- SSF-Dalen (Sweden-France Bilateral Collaboration) Program
- Chinese Scholarship Council
- INSERM
- CRITT Sante Bretagne (Region Bretagne)
- ANR Blanche of the French Government
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Domain V of the 23S/25S/28S rRNA of the large ribosomal subunit constitutes the active center for the protein folding activity of the ribosome (PFAR). Using in vitro transcribed domain V rRNAs from Escherichia coli and Saccharomyces cerevisiae as the folding modulators and human carbonic anhydrase as a model protein, we demonstrate that PFAR is conserved from prokaryotes to eukaryotes. It was shown previously that 6-aminophenanthridine (6AP), an antiprion compound, inhibits PFAR. Here, using UV cross-linking followed by primer extension, we show that the protein substrates and 6AP interact with a common set of nucleotides on domain V of 23S rRNA. Mutations at the interaction sites decreased PFAR and resulted in loss or change of the binding pattern for both the protein substrates and 6AP. Moreover, kinetic analysis of human carbonic anhydrase refolding showed that 6AP decreased the yield of the refolded protein but did not affect the rate of refolding. Thus, we conclude that 6AP competitively occludes the protein substrates from binding to rRNA and thereby inhibits PFAR. Finally, we propose a scheme clarifying the mechanism by which 6AP inhibits PFAR.
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