title: Distinct tRNA Accommodation Intermediates Observed on the Ribosome with the Antibiotics Hygromycin A and A201A.
abstract: The increase in multi-drug-resistant bacteria is limiting the effectiveness of currently approved antibiotics, leading to a renewed interest in antibiotics with distinct chemical scaffolds. We have solved the structures of the Thermus thermophilus 70S ribosome with A-, P-, and E-site tRNAs bound and in complex with either the aminocyclitol-containing antibiotic hygromycin A (HygA) or the nucleoside antibiotic A201A. Both antibiotics bind at the peptidyl transferase center and sterically occlude the CCA-end of the A-tRNA from entering the A site of the peptidyl transferase center. Single-molecule Förster resonance energy transfer (smFRET) experiments reveal that HygA and A201A specifically interfere with full accommodation of the A-tRNA, leading to the presence of tRNA accommodation intermediates and thereby inhibiting peptide bond formation. Thus, our results provide not only insight into the mechanism of action of HygA and A201A, but also into the fundamental process of tRNA accommodation during protein synthesis.
authors.affiliation: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
authors.forename: Yury S
authors.initials: YS
authors.lastname: Polikanov
authors.affiliation: Gene Center and Department for Biochemistry, University of Munich, Feodor-Lynenstr. 25, 81377 Munich, Germany.
authors.forename: Agata L
authors.initials: AL
authors.lastname: Starosta
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University, New York, NY 10065, USA.
authors.forename: Manuel F
authors.initials: MF
authors.lastname: Juette
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University, New York, NY 10065, USA.
authors.forename: Roger B
authors.initials: RB
authors.lastname: Altman
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University, New York, NY 10065, USA.
authors.forename: Daniel S
authors.initials: DS
authors.lastname: Terry
authors.affiliation: Department of Chemistry, Portland State University, Portland, OR 97207, USA.
authors.forename: Wanli
authors.initials: W
authors.lastname: Lu
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University, New York, NY 10065, USA.
authors.forename: Benjamin J
authors.initials: BJ
authors.lastname: Burnett
authors.affiliation: Department of Biochemistry, School of Medicine, University of Patras, 26500 Patras, Greece.
authors.forename: George
authors.initials: G
authors.lastname: Dinos
authors.affiliation: Department of Chemistry, Portland State University, Portland, OR 97207, USA.
authors.forename: Kevin A
authors.initials: KA
authors.lastname: Reynolds
authors.affiliation: Department of Physiology and Biophysics, Weill Medical College of Cornell University, New York, NY 10065, USA; Tri-Institutional Training Program in Chemical Biology, New York, NY 10065, USA. Electronic address: scb2005@med.cornell.edu.
authors.forename: Scott C
authors.initials: SC
authors.lastname: Blanchard
authors.affiliation: Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 06520, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA. Electronic address: thomas.steitz@yale.edu.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Steitz
authors.affiliation: Gene Center and Department for Biochemistry, University of Munich, Feodor-Lynenstr. 25, 81377 Munich, Germany; Center for integrated Protein Science Munich (CiPSM), University of Munich, Feodor-Lynenstr. 25, 81377 Munich, Germany. Electronic address: wilson@lmb.uni-muenchen.de.
authors.forename: Daniel N
authors.initials: DN
authors.lastname: Wilson
mesh_terms: D000617:Aminoglycosides / Q000737:chemistry* / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000494:pharmacology; D002934:Cinnamates / Q000737:chemistry* / Q000494:pharmacology; D018360:Crystallography, X-Ray; D052978:Disk Diffusion Antimicrobial Tests; D024881:Drug Resistance, Bacterial; D004926:Escherichia coli / Q000187:drug effects; D006860:Hydrogen Bonding; D006921:Hygromycin B / Q000031:analogs & derivatives* / Q000737:chemistry / Q000494:pharmacology; D008958:Models, Molecular; D011487:Protein Conformation; D012343:RNA, Transfer / Q000737:chemistry*; D054681:Ribosome Subunits, Large, Bacterial / Q000737:chemistry*; D054680:Ribosome Subunits, Small, Bacterial / Q000737:chemistry*; D016963:Thermus thermophilus
chemical_list: D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D002934:Cinnamates; C046076:A 201A; D006921:Hygromycin B; C026273:hygromycin A; D012343:RNA, Transfer