7N4E

Escherichia coli sigma 70-dependent paused transcription elongation complex

7N4E の概要

• エントリーDOI: 10.2210/pdb7n4e/pdb
• EMDBエントリー: 24148
• 分子名称: DNA (61-MER), MAGNESIUM ION, DNA-directed RNA polymerase subunit alpha, ... (10 entities in total)
• 機能のキーワード: elongation, pausing, sigma 70, transcription complex, dna scrunching, transferase-dna-rna complex, transferase/dna/rna
• 由来する生物種: Escherichia coli; 詳細
• タンパク質・核酸の鎖数: 9
• 化学式量合計: 501403.34

構造登録者

Molodtsov, V.,Su, M.,Ebright, R.H. (登録日: 2021-06-04, 公開日: 2022-06-15, 最終更新日: 2024-05-29)

主引用文献

Pukhrambam, C.,Molodtsov, V.,Kooshkbaghi, M.,Tareen, A.,Vu, H.,Skalenko, K.S.,Su, M.,Yin, Z.,Winkelman, J.T.,Kinney, J.B.,Ebright, R.H.,Nickels, B.E.
Structural and mechanistic basis of sigma-dependent transcriptional pausing.
Proc.Natl.Acad.Sci.USA, 119:e2201301119-e2201301119, 2022

PubMed Abstract: In σ-dependent transcriptional pausing, the transcription initiation factor σ, translocating with RNA polymerase (RNAP), makes sequence-specific protein–DNA interactions with a promoter-like sequence element in the transcribed region, inducing pausing. It has been proposed that, in σ-dependent pausing, the RNAP active center can access off-pathway “backtracked” states that are substrates for the transcript-cleavage factors of the Gre family and on-pathway “scrunched” states that mediate pause escape. Here, using site-specific protein–DNA photocrosslinking to define positions of the RNAP trailing and leading edges and of σ relative to DNA at the λPR′ promoter, we show directly that σ-dependent pausing in the absence of GreB in vitro predominantly involves a state backtracked by 2–4 bp, and σ-dependent pausing in the presence of GreB in vitro and in vivo predominantly involves a state scrunched by 2–3 bp. Analogous experiments with a library of 47 (∼16,000) transcribed-region sequences show that the state scrunched by 2–3 bp—and only that state—is associated with the consensus sequence, T−3N−2Y−1G+1, (where −1 corresponds to the position of the RNA 3′ end), which is identical to the consensus for pausing in initial transcription and which is related to the consensus for pausing in transcription elongation. Experiments with heteroduplex templates show that sequence information at position T−3 resides in the DNA nontemplate strand. A cryoelectron microscopy structure of a complex engaged in σ-dependent pausing reveals positions of DNA scrunching on the DNA nontemplate and template strands and suggests that position T−3 of the consensus sequence exerts its effects by facilitating scrunching.

PubMed: 35653571
DOI: 10.1073/pnas.2201301119

実験手法

ELECTRON MICROSCOPY (3.8 Å)