title: The antiprion compound 6-aminophenanthridine inhibits the protein folding activity of the ribosome by direct competition.
abstract: 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.
authors.affiliation: Department of Cell and Molecular Biology, Uppsala University, 75124 Uppsala, Sweden.
authors.forename: Yanhong
authors.initials: Y
authors.lastname: Pang
authors.forename: Sriram
authors.initials: S
authors.lastname: Kurella
authors.forename: Cécile
authors.initials: C
authors.lastname: Voisset
authors.forename: Dibyendu
authors.initials: D
authors.lastname: Samanta
authors.forename: Debapriya
authors.initials: D
authors.lastname: Banerjee
authors.forename: Ariane
authors.initials: A
authors.lastname: Schabe
authors.forename: Chanchal
authors.initials: C
authors.lastname: Das Gupta
authors.forename: Hervé
authors.initials: H
authors.lastname: Galons
authors.forename: Marc
authors.initials: M
authors.lastname: Blondel
authors.forename: Suparna
authors.initials: S
authors.lastname: Sanyal
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D001667:Binding, Competitive; D002256:Carbonic Anhydrases / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D018832:Molecular Chaperones / Q000737:chemistry; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D009690:Nucleic Acid Conformation; D010617:Phenanthridines / Q000494:pharmacology*; D011328:Prions / Q000737:chemistry*; D011485:Protein Binding; D011489:Protein Denaturation; D017510:Protein Folding / Q000187:drug effects*; D054730:Protein Interaction Domains and Motifs; D012335:RNA, Ribosomal / Q000737:chemistry; D012270:Ribosomes / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid
chemical_list: C549408:6-aminophenanthridine; D018832:Molecular Chaperones; D010617:Phenanthridines; D011328:Prions; D012335:RNA, Ribosomal; D002256:Carbonic Anhydrases
keywords: 6AP; Molecular Chaperone; Prions; Protein Folding; Ribosomal RNA (rRNA); Ribosomes