Dynamisches Zusammenspiel zwischen Chloramphenicol/Linezolid und dem translatierenden Ribosom
Zusammenfassung der Projektergebnisse
The translational machinery is a major target within the cell for antibiotics. Many clinically used classes of antibiotics inhibit translation by binding to the ribosome. Despite the potency of many of these drug classes, antibiotic resistance among clinically relevant pathogens is an increasing problem and thus the need for new antibiotics is more urgent than ever before. Here we have provided insight into the mechanism of action of a diverse range of antibiotic classes, including the third generation glycylcycline tigecycline, the orthosomycins evernimicin and avilamycin, the macrolide erythromycin and the antimicrobial apidaecin peptide. In addition, we have provided insight into how bacteria obtain resistance to tetracyclines using the ribosome protection protein TetM as well as to macrolides by inducing expression of resistance methyltransferase by utilizing drug- and polypeptide-dependent (ErmBL and ErmCL) stalling peptides. We are currently completing our studies and preparing manuscripts on the analogous Cat86L and CmlA stalling peptides that respond to chloramphenicols and oxazolidinones. Such stalling systems are not just utilized for antibiotic resistance but also for regulation of other cellular pathways and here we have provided structural insight into the mechanism by which the MifM and VemP stall the ribosome to regulate expression of YidC2 and SecDF2, respectively. MifM stalling is species-specific and occurs in Bacillus subtilis but not in E. coli. Thus, the development of the B. subtilis translation system and preparation of MifM-stalled ribosomes allowed us to also investigate the binding of the Alu-containing SRP at the tunnel exit of the ribosome. Studies into the folding of protein domains on the ribosome are still in progress and additional complexes will be investigated to attempt to catch and visualize stable intermediates at the tunnel exit. One of the most successful and collaborative projects within the FOR1805 addressed the mechanism of action of how polyproline stretches lead to translational stalling and how elongation EF-P and eIF-5A relieve this inhibition. In particular, illustrating how the novel post-translational modifications on these factors stabilizes the P-site tRNA, particularly via interactions between its modification and the CCA end, thereby enforcing an alternative conformation of the polyproline-containing nascent chain, which allows a favorable substrate geometry for peptide bond formation.
Projektbezogene Publikationen (Auswahl)
-
Structural basis for TetM-mediated tetracycline resistance. Proc Natl Acad Sci USA. 109(42):16900-5 (2012)
Dönhöfer A, Franckenberg S, Wickles S, Berninghausen O, Beckmann R, Wilson DN
-
Structural basis for potent inhibitory activity of the antibiotic tigecycline during protein synthesis. Proc Natl Acad Sci USA. 110(10):3812-6 (2013)
Jenner L, Starosta AL, Terry DS, Mikolajka A, Filonava L, Yusupov M, Blanchard SC, Wilson DN, Yusupova G
-
Cryo-EM structure of the tetracycline resistance protein TetM in complex with a translating ribosome at 3.9-Å resolution. Proc Natl Acad Sci USA. 112(17):5401-6 (2015)
Arenz S, Nguyen F, Beckmann R, Wilson DN
-
Entropic Contribution of Elongation Factor P to Proline Positioning at the Catalytic Center of the Ribosome. J Am Chem Soc. 137:12997-3006 (2015)
Doerfel LK, Wohlgemuth I, Kubyshkin V, Starosta AL, Wilson DN, Budisa N, Rodnina MV
-
A combined cryo-EM and molecular dynamics approach reveals the mechanism of ErmBL-mediated translation arrest. Nat. Commun. 7:12026 (2016)
Arenz S, Bock LV, Graf M, Innis CA, Beckmann R, Grubmüller H, Vaiana AC, Wilson DN
-
An antimicrobial peptide that inhibits translation by trapping release factors on the ribosome. Nat Struct Mol Biol. 24:752-757 (2017)
Florin T, Maracci C, Graf M, Karki P, Klepacki D, Berninghausen O, Beckmann R, Vázquez-Laslop N, Wilson DN, Rodnina MV, Mankin AS
-
Structural basis for ArfA-RF2-mediated translation termination on mRNAs lacking stop codons. Nature. 541(7638):546-549 (2017)
Huter P, Müller C, Beckert B, Arenz S, Berninghausen O, Beckmann R, Wilson DN
-
Structural Basis for Polyproline-Mediated Ribosome Stalling and Rescue by the Translation Elongation Factor EF-P. Molecular Cell. 68(3):515-527 (2017)
Huter P, Arenz S, Bock LV, Graf M, Frister JO, Heuer A, Peil L, Starosta AL, Wohlgemuth I, Peske F, Nováček J, Berninghausen O, Grubmüller H, Tenson T, Beckmann R, Rodnina MV, Vaiana AC, Wilson DN
-
Structure of a hibernating 100S ribosome reveals an inactive conformation of the ribosomal protein S1. Nat Microbiol. 3(10):1115-1121 (2018)
Beckert B, Turk M, Czech A, Berninghausen O, Beckmann R, Ignatova Z, Plitzko JM, Wilson DN
-
Visualization of translation termination intermediates trapped by the Apidaecin 137 peptide during RF3-mediated recycling of RF1. Nat Commun. 9(1):3053 (2018)
Graf M, Huter P, Maracci C, Peterek M, Rodnina MV, Wilson DN
