Structures of an RNA polymerase promoter melting intermediate elucidate DNA unwinding

Nature. 2019 Jan;565(7739):382-385. doi: 10.1038/s41586-018-0840-5. Epub 2019 Jan 9.

Abstract

A key regulated step of transcription is promoter melting by RNA polymerase (RNAP) to form the open promoter complex1-3. To generate the open complex, the conserved catalytic core of the RNAP combines with initiation factors to locate promoter DNA, unwind 12-14 base pairs of the DNA duplex and load the template-strand DNA into the RNAP active site. Formation of the open complex is a multi-step process during which transient intermediates of unknown structure are formed4-6. Here we present cryo-electron microscopy structures of bacterial RNAP-promoter DNA complexes, including structures of partially melted intermediates. The structures show that late steps of promoter melting occur within the RNAP cleft, delineate key roles for fork-loop 2 and switch 2-universal structural features of RNAP-in restricting access of DNA to the RNAP active site, and explain why clamp opening is required to allow entry of single-stranded template DNA into the active site. The key roles of fork-loop 2 and switch 2 suggest a common mechanism for late steps in promoter DNA opening to enable gene expression across all domains of life.

Publication types

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

MeSH terms

  • Bacterial Proteins / metabolism
  • Base Sequence
  • Catalytic Domain
  • Cryoelectron Microscopy*
  • DNA, Bacterial / chemistry*
  • DNA, Bacterial / metabolism
  • DNA, Bacterial / ultrastructure*
  • DNA-Directed RNA Polymerases / metabolism*
  • Enzyme Stability / drug effects
  • Escherichia coli / enzymology
  • Lactones / pharmacology
  • Models, Molecular
  • Mycobacterium tuberculosis / enzymology*
  • Mycobacterium tuberculosis / metabolism
  • Nucleic Acid Conformation*
  • Nucleic Acid Denaturation
  • Promoter Regions, Genetic*
  • Protein Binding
  • Thermodynamics
  • Transcription Initiation, Genetic / drug effects

Substances

  • Bacterial Proteins
  • DNA, Bacterial
  • Lactones
  • corallopyronin A
  • myxopyronin A
  • DNA-Directed RNA Polymerases