Structural Basis of Transcription Inhibition by Fidaxomicin (Lipiarmycin A3)

Mol Cell. 2018 Apr 5;70(1):60-71.e15. doi: 10.1016/j.molcel.2018.02.026. Epub 2018 Mar 29.

Abstract

Fidaxomicin is an antibacterial drug in clinical use for treatment of Clostridium difficile diarrhea. The active ingredient of fidaxomicin, lipiarmycin A3 (Lpm), functions by inhibiting bacterial RNA polymerase (RNAP). Here we report a cryo-EM structure of Mycobacterium tuberculosis RNAP holoenzyme in complex with Lpm at 3.5-Å resolution. The structure shows that Lpm binds at the base of the RNAP "clamp." The structure exhibits an open conformation of the RNAP clamp, suggesting that Lpm traps an open-clamp state. Single-molecule fluorescence resonance energy transfer experiments confirm that Lpm traps an open-clamp state and define effects of Lpm on clamp dynamics. We suggest that Lpm inhibits transcription by trapping an open-clamp state, preventing simultaneous interaction with promoter -10 and -35 elements. The results account for the absence of cross-resistance between Lpm and other RNAP inhibitors, account for structure-activity relationships of Lpm derivatives, and enable structure-based design of improved Lpm derivatives.

Keywords: Mycobacterium tuberculosis; RNA polymerase; RNA polymerase clamp; RNA polymerase inhibitor; RNA polymerase switch region; antibiotic; cryo-electron microscopy; fidaxomicin; lipiarmycin; single-molecule fluorescence resonance energy transfer.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / chemistry
  • Anti-Bacterial Agents / metabolism
  • Anti-Bacterial Agents / pharmacology*
  • Bacterial Proteins / antagonists & inhibitors*
  • Bacterial Proteins / metabolism
  • Bacterial Proteins / ultrastructure
  • Binding Sites
  • Cryoelectron Microscopy
  • DNA-Directed RNA Polymerases / antagonists & inhibitors*
  • DNA-Directed RNA Polymerases / metabolism
  • DNA-Directed RNA Polymerases / ultrastructure
  • Drug Design
  • Drug Resistance, Bacterial / genetics
  • Escherichia coli / drug effects*
  • Escherichia coli / enzymology
  • Escherichia coli / genetics
  • Escherichia coli / ultrastructure
  • Fidaxomicin / chemistry
  • Fidaxomicin / metabolism
  • Fidaxomicin / pharmacology*
  • Fluorescence Resonance Energy Transfer
  • Gene Expression Regulation, Bacterial / drug effects
  • Models, Molecular
  • Mutation
  • Mycobacterium tuberculosis / drug effects*
  • Mycobacterium tuberculosis / enzymology
  • Mycobacterium tuberculosis / genetics
  • Mycobacterium tuberculosis / ultrastructure
  • Protein Binding
  • Protein Conformation
  • Single Molecule Imaging
  • Staphylococcus aureus / drug effects
  • Staphylococcus aureus / enzymology
  • Staphylococcus aureus / genetics
  • Structure-Activity Relationship
  • Transcription, Genetic / drug effects*

Substances

  • Anti-Bacterial Agents
  • Bacterial Proteins
  • DNA-Directed RNA Polymerases
  • Fidaxomicin