STRUCTURAL BASIS OF TRANSCRIPTION INHIBITION BY FIDAXOMICIN (LIPIARMYCIN A3)
| Autor: | Yi Jiang, Kalyan Das, Diego Duchi, Aashish Srivastava, Achillefs N. Kapanidis, Dongye Wang, Stefano Donadio, Ruiheng Yin, Haibo Zhang, Edward T. Eng, Changsheng Zhang, Abhishek Mazumder, Elena V. Sineva, Yon W. Ebright, Richard Y. Ebright, Dennis Thomas, Sukhendu Mandal, Matthew Gigliotti, David Degen, Wei Lin, Yu Liu, Richard H. Ebright, Zhening Zhang |
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| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Models Molecular Transcription Genetic Protein Conformation medicine.disease_cause Lipiarmycin A3 chemistry.chemical_compound Transcription (biology) RNA polymerase Fluorescence Resonance Energy Transfer Fidaxomicin Polymerase 0303 health sciences biology Chemistry DNA-Directed RNA Polymerases Single Molecule Imaging Anti-Bacterial Agents medicine.drug Protein Binding Staphylococcus aureus 030106 microbiology Article 03 medical and health sciences Structure-Activity Relationship Bacterial Proteins Drug Resistance Bacterial medicine Escherichia coli Binding site Molecular Biology 030304 developmental biology Binding Sites 030306 microbiology Cryoelectron Microscopy RNA Cell Biology Gene Expression Regulation Bacterial Mycobacterium tuberculosis Clostridium difficile diarrhea enzymes and coenzymes (carbohydrates) 030104 developmental biology Förster resonance energy transfer RNA Polymerase Inhibitor Drug Design Mutation biology.protein Biophysics bacteria |
| Popis: | Fidaxomicin is an antibacterial drug in clinical use in treatment ofClostridium difficilediarrhea1–2. The active pharmaceutical ingredient of fidaxomicin, lipiarmycin A3 (Lpm)1–4, is a macrocyclic antibiotic with bactericidal activity against Gram-positive bacteria and efflux-deficient strains of Gram-negative bacteria1–2, 5. Lpm functions by inhibiting bacterial RNA polymerase (RNAP)6–8. Lpm exhibits no cross-resistance with the classic RNAP inhibitor rifampin (Rif)7, 9and inhibits transcription initiation at an earlier step than Rif8–11, suggesting that the binding site and mechanism of Lpm differ from those of Rif. Efforts spanning a decade to obtain a crystal structure of RNAP in complex with Lpm have been unsuccessful. Here, we report a cryo-EM12–13structure ofMycobacterium tuberculosisRNAP holoenzyme in complex with Lpm at 3.5 Å resolution. The structure shows that Lpm binds at the base of the RNAP “clamp,” interacting with the RNAP switch region and the RNAP RNA exit channel. The binding site on RNAP for Lpm does not overlap the binding sites for other RNAP inhibitors, accounting for the absence of cross-resistance of Lpm with other RNAP inhibitors. The structure exhibits an open conformation of the RNAP clamp, with the RNAP clamp swung outward by ~17° relative to its position in catalytically competent RNAP-promoter transcription initiation complexes, suggesting that Lpm traps an open-clamp conformational state. Single-molecule fluorescence resonance energy transfer14experiments confirm that Lpm traps an open-clamp conformational state and define effects of Lpm on clamp opening and closing dynamics. We propose that Lpm inhibits transcription initiation by trapping an open-clamp conformational state, thereby preventing simultaneous engagement of transcription initiation factor σ regions 2 and 4 with promoter -10 and -35 elements. The results provide information essential to understanding the mode of action of Lpm, account for structure-activity relationships of known Lpm analogs, and suggest modifications to Lpm that could yield new, improved Lpm analogs. |
| Databáze: | OpenAIRE |
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