4.8 Article

RNA polymerase clamp conformational dynamics: long-lived states and modulation by crowding, cations, and nonspecific DNA binding

Journal

NUCLEIC ACIDS RESEARCH
Volume 49, Issue 5, Pages 2790-2802

Publisher

OXFORD UNIV PRESS
DOI: 10.1093/nar/gkab074

Keywords

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Funding

  1. Wellcome Trust [110164/Z/15/Z]
  2. NIH [GM041376]
  3. Wellcome Trust [110164/Z/15/Z] Funding Source: Wellcome Trust

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The study reveals that the RNA polymerase clamp can transition between different conformational states and exhibit varying dynamics under different environmental conditions. These findings suggest potential regulatory checkpoints that could impact clamp dynamics and transcription processes.
The RNA polymerase (RNAP) clamp, a mobile structural element conserved in RNAP from all domains of life, has been proposed to play critical roles at different stages of transcription. In previous work, we demonstrated using single-molecule Forster resonance energy transfer (smFRET) that RNAP clamp interconvert between three short-lived conformational states (lifetimes similar to 0.3-0.6 s), that the clamp can be locked into any one of these states by small molecules, and that the clamp stays closed during initial transcription and elongation. Here, we extend these studies to obtain a comprehensive understanding of clamp dynamics under conditions RNAP may encounter in living cells. We find that the RNAP clamp can populate long-lived conformational states (lifetimes > 1.0 s) and can switch between these long-lived states and the previously observed short-lived states. In addition, we find that clamp motions are increased in the presence of molecular crowding, are unchanged in the presence of elevated monovalent-cation concentrations, and are reduced in the presence of elevated divalent-cation concentrations. Finally, we find that RNAP bound to non-specific DNA predominantly exhibits a closed clamp conformation. Our results raise the possibility of additional regulatory checkpoints that could affect clamp dynamics and consequently could affect transcription and transcriptional regulation.

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