8P2I

Cryo-EM structure of Pyrococcus furiosus apo form RNA polymerase contracted clamp conformation with Spt4/5

Summary for 8P2I

• Entry DOI: 10.2210/pdb8p2i/pdb
• EMDB information: 17366
• Descriptor: DNA-directed RNA polymerase subunit Rpo1N, DNA-directed RNA polymerase subunit Rpo10, DNA-directed RNA polymerase subunit Rpo12, ... (15 entities in total)
• Functional Keywords: rna, polymerase, elongation, complex, pyrococcus furiosus, archaea, transcription, dna
• Biological source: Pyrococcus furiosus DSM 3638; More
• Total number of polymer chains: 13
• Total formula weight: 407210.92

Authors

Tarau, D.M.,Reichelt, R.,Heiss, F.B.,Pilsl, M.,Hausner, W.,Engel, C.,Grohmann, D. (deposition date: 2023-05-16, release date: 2024-04-24, Last modification date: 2025-07-09)

Primary citation

Tarau, D.,Grunberger, F.,Pilsl, M.,Reichelt, R.,Heiss, F.,Konig, S.,Urlaub, H.,Hausner, W.,Engel, C.,Grohmann, D.
Structural basis of archaeal RNA polymerase transcription elongation and Spt4/5 recruitment.
Nucleic Acids Res., 52:6017-6035, 2024

PubMed Abstract: Archaeal transcription is carried out by a multi-subunit RNA polymerase (RNAP) that is highly homologous in structure and function to eukaryotic RNAP II. Among the set of basal transcription factors, only Spt5 is found in all domains of life, but Spt5 has been shaped during evolution, which is also reflected in the heterodimerization of Spt5 with Spt4 in Archaea and Eukaryotes. To unravel the mechanistic basis of Spt4/5 function in Archaea, we performed structure-function analyses using the archaeal transcriptional machinery of Pyrococcus furiosus (Pfu). We report single-particle cryo-electron microscopy reconstructions of apo RNAP and the archaeal elongation complex (EC) in the absence and presence of Spt4/5. Surprisingly, Pfu Spt4/5 also binds the RNAP in the absence of nucleic acids in a distinct super-contracted conformation. We show that the RNAP clamp/stalk module exhibits conformational flexibility in the apo state of RNAP and that the enzyme contracts upon EC formation or Spt4/5 engagement. We furthermore identified a contact of the Spt5-NGN domain with the DNA duplex that stabilizes the upstream boundary of the transcription bubble and impacts Spt4/5 activity in vitro. This study, therefore, provides the structural basis for Spt4/5 function in archaeal transcription and reveals a potential role beyond the well-described support of elongation.

PubMed: 38709902
DOI: 10.1093/nar/gkae282

Experimental method

ELECTRON MICROSCOPY (3.4 Å)