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Structure paper

TitleStructural basis for multi-subunit DNA-dependent RNA polymerase catalytic activity.
Journal, issue, pagesMol Cell, Year 2026
Publish dateApr 30, 2026
AuthorsAndreas U Mueller / Seth A Darst /
PubMed AbstractMulti-subunit DNA-dependent RNA polymerase (RNAP) is the central enzyme of transcription, yet its catalytic mechanism remains obscure because high-resolution structures of intermediates with native ...Multi-subunit DNA-dependent RNA polymerase (RNAP) is the central enzyme of transcription, yet its catalytic mechanism remains obscure because high-resolution structures of intermediates with native substrates are not available. We visualized E. coli RNAP on a promoter DNA template with nucleoside triphosphate substrates engaged in active RNA synthesis by cryo-electron microscopy. From this heterogeneous mixture, we determined five high-resolution structures of initial transcribing complexes, including a true Michaelis complex (MC) and a post-catalytic product complex (PC). The MC reveals key conformational transitions during catalysis. Waters in the MC and PC structures show striking overlap with those in corresponding S. cerevisiae RNA polymerase II (RNAPII) structures (see related paper by Li et al.), pointing to functional importance. Together, these results establish that RNAP catalyzes nucleotidyl transfer through a positional (entropic) mechanism, revealing structural determinants of the first step of gene expression.
External linksMol Cell / PubMed:42066755
MethodsEM (single particle)
Resolution2.5 - 3.1 Å
Structure data

73122

9ymu

EMDB-73122, PDB-9ymu:
De novo initial transcribing RNA polymerase with 2-mer RNA and bound CTP / Michaelis complex (RPitc2+CTP)
Method: EM (single particle) / Resolution: 2.7 Å

73123

9ymv

EMDB-73123, PDB-9ymv:
De novo initial transcribing RNA polymerase with pretranslocated 3-mer RNA / product complex (RPitc3)
Method: EM (single particle) / Resolution: 2.6 Å

73124

9ymw

EMDB-73124: de novo initial transcribing RNA polymerase with 5-mer RNA CCAUC (RPitc5a)
PDB-9ymw: De novo initial transcribing RNA polymerase with 5-mer RNA CCAUC (RPitc5a)
Method: EM (single particle) / Resolution: 2.7 Å

73125

9ymx

EMDB-73125, PDB-9ymx:
De novo initial transcribing RNA polymerase with 5-mer RNA AUCUA (RPitc5b)
Method: EM (single particle) / Resolution: 2.7 Å

73126

9ymy

EMDB-73126, PDB-9ymy:
De novo initial transcribing RNA polymerase with 9-mer RNA (RPitc9)
Method: EM (single particle) / Resolution: 3.1 Å

EMDB-73127: De novo initial transcribing RNA polymerase on T7A1 promoter (weak downstream)
Method: EM (single particle) / Resolution: 3.0 Å

EMDB-73128: De novo initial transcribing RNA polymerase on T7A1 promoter (empty channel)
Method: EM (single particle) / Resolution: 2.5 Å

EMDB-73130: RNA polymerase open complex on T7A1 promoter (RPo)
Method: EM (single particle) / Resolution: 2.8 Å

73131

9yn0

EMDB-73131, PDB-9yn0:
De novo initial transcribing RNA polymerase with sigma 70 region 1.1 bound (RPitc-s70_1.1)
Method: EM (single particle) / Resolution: 2.9 Å

Chemicals

ChemComp-MG:
Unknown entry

ChemComp-ZN:
Unknown entry

ChemComp-CTP:
CYTIDINE-5'-TRIPHOSPHATE

ChemComp-HOH:
WATER

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

ChemComp-POP:
PYROPHOSPHATE 2-

Source
  • escherichia coli (E. coli)
  • escherichia phage t7 (virus)
KeywordsTRANSCRIPTION / RNA polymerase / initial transcribing complex / substrate-bound / nucleotide addition / pre-catalytic / post-catalytic / promoter escape / transcription initiation / sigma 70 region 1.1

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