National Natural Science Foundation of China (NSFC)
31822001
China
Citation
Journal: Nature / Year: 2023 Title: Structural basis for intrinsic transcription termination. Authors: Linlin You / Expery O Omollo / Chengzhi Yu / Rachel A Mooney / Jing Shi / Liqiang Shen / Xiaoxian Wu / Aijia Wen / Dingwei He / Yuan Zeng / Yu Feng / Robert Landick / Yu Zhang / Abstract: Efficient and accurate termination is required for gene transcription in all living organisms. Cellular RNA polymerases in both bacteria and eukaryotes can terminate their transcription through a ...Efficient and accurate termination is required for gene transcription in all living organisms. Cellular RNA polymerases in both bacteria and eukaryotes can terminate their transcription through a factor-independent termination pathway-called intrinsic termination transcription in bacteria-in which RNA polymerase recognizes terminator sequences, stops nucleotide addition and releases nascent RNA spontaneously. Here we report a set of single-particle cryo-electron microscopy structures of Escherichia coli transcription intrinsic termination complexes representing key intermediate states of the event. The structures show how RNA polymerase pauses at terminator sequences, how the terminator RNA hairpin folds inside RNA polymerase, and how RNA polymerase rewinds the transcription bubble to release RNA and then DNA. These macromolecular snapshots define a structural mechanism for bacterial intrinsic termination and a pathway for RNA release and DNA collapse that is relevant for factor-independent termination by all RNA polymerases.
Name: RNA (5'-R(*CP*CP*UP*GP*CP*GP*AP*CP*GP*C)-3') / type: rna / ID: 8 / Number of copies: 1
Source (natural)
Organism: synthetic construct (others)
Molecular weight
Theoretical: 3.151941 KDa
Sequence
String:
CCUGCGACGC
+
Macromolecule #9: ZINC ION
Macromolecule
Name: ZINC ION / type: ligand / ID: 9 / Number of copies: 2 / Formula: ZN
Molecular weight
Theoretical: 65.409 Da
+
Macromolecule #10: MAGNESIUM ION
Macromolecule
Name: MAGNESIUM ION / type: ligand / ID: 10 / Number of copies: 1 / Formula: MG
Molecular weight
Theoretical: 24.305 Da
-
Experimental details
-
Structure determination
Method
cryo EM
Processing
single particle reconstruction
Aggregation state
particle
-
Sample preparation
Concentration
12 mg/mL
Buffer
pH: 7.5
Grid
Model: C-flat-1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 40 / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: OTHER
Vitrification
Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV
-
Electron microscopy
Microscope
FEI TITAN KRIOS
Image recording
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number real images: 3888 / Average exposure time: 5.85 sec. / Average electron dose: 58.5 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
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