title: The antibiotics dityromycin and GE82832 bind protein S12 and block EF-G-catalyzed translocation.
abstract: The translocation of mRNA and tRNA through the ribosome is catalyzed by elongation factor G (EF-G), a universally conserved guanosine triphosphate hydrolase (GTPase). The mechanism by which the closely related decapeptide antibiotics dityromycin and GE82832 inhibit EF-G-catalyzed translocation is elucidated in this study. Using crystallographic and biochemical experiments, we demonstrate that these antibiotics bind to ribosomal protein S12 in solution alone as well as within the small ribosomal subunit, inducing long-range effects on the ribosomal head. The crystal structure of the antibiotic in complex with the 70S ribosome reveals that the binding involves conserved amino acid residues of S12 whose mutations result in in vitro and in vivo antibiotic resistance and loss of antibiotic binding. The data also suggest that GE82832/dityromycin inhibits EF-G-catalyzed translocation by disrupting a critical contact between EF-G and S12 that is required to stabilize the posttranslocational conformation of EF-G, thereby preventing the ribosome-EF-G complex from entering a conformation productive for translocation.
authors.affiliation: Department of Chemistry, Yale University, New Haven, CT 06511, USA.
authors.forename: David
authors.initials: D
authors.lastname: Bulkley
authors.affiliation: Laboratory of Genetics, Department of Biosciences and Biotechnology, University of Camerino, 62032 Camerino, Italy.
authors.forename: Letizia
authors.initials: L
authors.lastname: Brandi
authors.affiliation: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06511, USA; Howard Hughes Medical Institute, New Haven, CT 06511, USA.
authors.forename: Yury S
authors.initials: YS
authors.lastname: Polikanov
authors.affiliation: Laboratory of Genetics, Department of Biosciences and Biotechnology, University of Camerino, 62032 Camerino, Italy.
authors.forename: Attilio
authors.initials: A
authors.lastname: Fabbretti
authors.affiliation: School of Biological Sciences, University of Missouri-Kansas City, Kansas City, MO 64110, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: O'Connor
authors.affiliation: Laboratory of Genetics, Department of Biosciences and Biotechnology, University of Camerino, 62032 Camerino, Italy. Electronic address: claudio.gualerzi@unicam.it.
authors.forename: Claudio O
authors.initials: CO
authors.lastname: Gualerzi
authors.affiliation: Department of Chemistry, Yale University, New Haven, CT 06511, USA; Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06511, USA; Howard Hughes Medical Institute, New Haven, CT 06511, USA. Electronic address: thomas.steitz@yale.edu.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Steitz
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D001665:Binding Sites; D004926:Escherichia coli / Q000187:drug effects; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D010441:Peptide Chain Elongation, Translational / Q000187:drug effects*; D020652:Peptide Elongation Factor 2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D011485:Protein Binding; D016963:Thermus thermophilus / Q000187:drug effects
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; C512383:GE82832 peptide, Streptosporangium cinnabarinum; D020652:Peptide Elongation Factor 2; D010455:Peptides