Japan Agency for Medical Research and Development (AMED)
JP21am0101071
Japan
Japan Society for the Promotion of Science (JSPS)
21K06032
Japan
Japan Society for the Promotion of Science (JSPS)
JP18J00191
Japan
Citation
Journal: J Biol Chem / Year: 2021 Title: Minimal protein-only RNase P structure reveals insights into tRNA precursor recognition and catalysis. Authors: Takamasa Teramoto / Takeshi Koyasu / Naruhiko Adachi / Masato Kawasaki / Toshio Moriya / Tomoyuki Numata / Toshiya Senda / Yoshimitsu Kakuta / Abstract: Ribonuclease P (RNase P) is an endoribonuclease that catalyzes the processing of the 5' leader sequence of precursor tRNA (pre-tRNA). Ribonucleoprotein RNase P and protein-only RNase P (PRORP) in ...Ribonuclease P (RNase P) is an endoribonuclease that catalyzes the processing of the 5' leader sequence of precursor tRNA (pre-tRNA). Ribonucleoprotein RNase P and protein-only RNase P (PRORP) in eukaryotes have been extensively studied, but the mechanism by which a prokaryotic nuclease recognizes and cleaves pre-tRNA is unclear. To gain insights into this mechanism, we studied homologs of Aquifex RNase P (HARPs), thought to be enzymes of approximately 23 kDa comprising only this nuclease domain. We determined the cryo-EM structure of Aq880, the first identified HARP enzyme. The structure unexpectedly revealed that Aq880 consists of both the nuclease and protruding helical (PrH) domains. Aq880 monomers assemble into a dimer via the PrH domain. Six dimers form a dodecamer with a left-handed one-turn superhelical structure. The structure also revealed that the active site of Aq880 is analogous to that of eukaryotic PRORPs. The pre-tRNA docking model demonstrated that 5' processing of pre-tRNAs is achieved by two adjacent dimers within the dodecamer. One dimer is responsible for catalysis, and the PrH domains of the other dimer are responsible for pre-tRNA elbow recognition. Our study suggests that HARPs measure an invariant distance from the pre-tRNA elbow to cleave the 5' leader sequence, which is analogous to the mechanism of eukaryotic PRORPs and the ribonucleoprotein RNase P. Collectively, these findings shed light on how different types of RNase P enzymes utilize the same pre-tRNA processing.
History
Deposition
Jun 16, 2021
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Header (metadata) release
Aug 11, 2021
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Map release
Aug 11, 2021
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Update
Feb 23, 2022
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Current status
Feb 23, 2022
Processing site: PDBj / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
EMPIAR-11072 (Title: Minimal protein-only RNase P structure reveals insights into tRNA precursor recognition and catalysis Data size: 2.0 TB Data #1: Minimal protein-only RNase P structure reveals insights into tRNA precursor recognition and catalysis [micrographs - multiframe])
Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Details: The grid was washed by acetone prior to use.
Vitrification
Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 291 K / Instrument: FEI VITROBOT MARK IV / Details: Blotting time was 5 seconds (blot force 20).
Details
This sample was mono-disperse.
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Electron microscopy
Microscope
TFS TALOS
Electron beam
Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Film or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 2370 / Average exposure time: 48.62 sec. / Average electron dose: 50.0 e/Å2
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Image processing
Particle selection
Number selected: 1486899
CTF correction
Software - Name: Gctf
Startup model
Type of model: OTHER Details: An ab initio model was generated using RELION3's own implementation of Stochastic Gradient Descent (SGD) algorithm and low-pass filtered to 60 A for use as an initial model for 3D classification.
Initial angle assignment
Type: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
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