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- PDB-7y17: Crystal structure of ribosomal ITS2 pre-rRNA processing complex f... -

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Basic information

Entry
Database: PDB / ID: 7y17
TitleCrystal structure of ribosomal ITS2 pre-rRNA processing complex from Cyberlindnera jadinii
Components
  • LAS1 protein
  • Polynucleotide 5'-hydroxyl-kinase GRC3
KeywordsRNA BINDING PROTEIN / RNA processing / Nuclease
Function / homology
Function and homology information


polynucleotide 5'-hydroxyl-kinase activity / Las1 complex / maturation of 5.8S rRNA / preribosome, large subunit precursor / RNA processing / maturation of LSU-rRNA / endonuclease activity / ATP binding
Similarity search - Function
Las1 / Las1-like / Polyribonucleotide 5'-hydroxyl-kinase Clp1, P-loop domain / Polyribonucleotide 5-hydroxyl-kinase Clp1/Grc3 / mRNA cleavage and polyadenylation factor CLP1 P-loop / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Polynucleotide 5'-hydroxyl-kinase GRC3 / LAS1 protein
Similarity search - Component
Biological speciesCyberlindnera jadinii (fungus)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.39 Å
AuthorsChen, J. / Liu, L.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32171286 China
CitationJournal: Elife / Year: 2024
Title: Structural and mechanistic insights into ribosomal ITS2 RNA processing by nuclease-kinase machinery.
Authors: Jiyun Chen / Hong Chen / Shanshan Li / Xiaofeng Lin / Rong Hu / Kaiming Zhang / Liang Liu /
Abstract: Precursor ribosomal RNA (pre-rRNA) processing is a key step in ribosome biosynthesis and involves numerous RNases. A HEPN (higher eukaryote and prokaryote nucleotide binding) nuclease Las1 and a ...Precursor ribosomal RNA (pre-rRNA) processing is a key step in ribosome biosynthesis and involves numerous RNases. A HEPN (higher eukaryote and prokaryote nucleotide binding) nuclease Las1 and a polynucleotide kinase Grc3 assemble into a tetramerase responsible for rRNA maturation. Here, we report the structures of full-length and Las1-Grc3 complexes, and Las1. The Las1-Grc3 structures show that the central coiled-coil domain of Las1 facilitates pre-rRNA binding and cleavage, while the Grc3 C-terminal loop motif directly binds to the HEPN active center of Las1 and regulates pre-rRNA cleavage. Structural comparison between Las1 and Las1-Grc3 complex exhibits that Grc3 binding induces conformational rearrangements of catalytic residues associated with HEPN nuclease activation. Biochemical assays identify that Las1 processes pre-rRNA at the two specific sites (C2 and C2'), which greatly facilitates rRNA maturation. Our structures and specific pre-rRNA cleavage findings provide crucial insights into the mechanism and pathway of pre-rRNA processing in ribosome biosynthesis.
History
DepositionJun 7, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jun 14, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 29, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Revision 1.2Jan 17, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: Polynucleotide 5'-hydroxyl-kinase GRC3
C: LAS1 protein
D: LAS1 protein
A: Polynucleotide 5'-hydroxyl-kinase GRC3
E: Polynucleotide 5'-hydroxyl-kinase GRC3
F: LAS1 protein


Theoretical massNumber of molelcules
Total (without water)357,0796
Polymers357,0796
Non-polymers00
Water6,954386
1
B: Polynucleotide 5'-hydroxyl-kinase GRC3
C: LAS1 protein
D: LAS1 protein
A: Polynucleotide 5'-hydroxyl-kinase GRC3
E: Polynucleotide 5'-hydroxyl-kinase GRC3


Theoretical massNumber of molelcules
Total (without water)307,6665
Polymers307,6665
Non-polymers00
Water905
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area13010 Å2
ΔGint-87 kcal/mol
Surface area77500 Å2
MethodPISA
2
F: LAS1 protein

F: LAS1 protein


Theoretical massNumber of molelcules
Total (without water)98,8272
Polymers98,8272
Non-polymers00
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation3_554-x,y,-z-1/21
Buried area11230 Å2
ΔGint-80 kcal/mol
Surface area81610 Å2
MethodPISA
Unit cell
Length a, b, c (Å)152.619, 240.021, 237.003
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number20
Space group name H-MC2221
Space group name HallC2c2
Symmetry operation#1: x,y,z
#2: x,-y,-z
#3: -x,y,-z+1/2
#4: -x,-y,z+1/2
#5: x+1/2,y+1/2,z
#6: x+1/2,-y+1/2,-z
#7: -x+1/2,y+1/2,-z+1/2
#8: -x+1/2,-y+1/2,z+1/2
Components on special symmetry positions
IDModelComponents
11C-531-

HOH

21D-545-

HOH

31D-552-

HOH

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Components

#1: Protein Polynucleotide 5'-hydroxyl-kinase GRC3 / Polynucleotide 5'-hydroxyl-kinase grc3


Mass: 69612.867 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cyberlindnera jadinii (fungus)
Strain: ATCC 18201 / CBS 1600 / BCRC 20928 / JCM 3617 / NBRC 0987 / NRRL Y-1542
Gene: GRC3, BN1211_2636, CYBJADRAFT_171070 / Production host: Escherichia coli K-12 (bacteria) / References: UniProt: A0A0H5C3P3
#2: Protein LAS1 protein


Mass: 49413.621 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Cyberlindnera jadinii (fungus)
Strain: ATCC 18201 / CBS 1600 / BCRC 20928 / JCM 3617 / NBRC 0987 / NRRL Y-1542
Gene: LAS1, BN1211_1791 / Production host: Escherichia coli K-12 (bacteria) / References: UniProt: A0A0H5CBH3
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 386 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 3.04 Å3/Da / Density % sol: 59.53 %
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop
Details: 0.1 M sodium phosphate, pH 7.5, 0.05 M NaCl, and 9% PEG 4000

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRF / Beamline: BL18U1 / Wavelength: 0.9793 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jan 12, 2020
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9793 Å / Relative weight: 1
ReflectionResolution: 3.23→50 Å / Num. obs: 41321 / % possible obs: 99.9 % / Redundancy: 7.5 % / Biso Wilson estimate: 71.33 Å2 / Rpim(I) all: 0.135 / Net I/σ(I): 4.3
Reflection shellResolution: 3.23→3.29 Å / Num. unique obs: 3403 / Rpim(I) all: 0.412

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Processing

Software
NameVersionClassification
PHENIX1.17.1_3660refinement
HKL-30007.21data scaling
HKL-30007.21data reduction
Coot0.7.2.1model building
PHENIX1.17.1_3660phasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 6OF4

6of4


Resolution: 3.39→21.41 Å / SU ML: 0.6081 / Cross valid method: FREE R-VALUE / σ(F): 1.41 / Phase error: 31.8318
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.3319 2096 5.07 %
Rwork0.3087 39225 -
obs0.3099 41321 68.35 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 124.13 Å2
Refinement stepCycle: LAST / Resolution: 3.39→21.41 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms18732 0 0 386 19118
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.011419096
X-RAY DIFFRACTIONf_angle_d1.786626006
X-RAY DIFFRACTIONf_chiral_restr0.26183111
X-RAY DIFFRACTIONf_plane_restr0.00763295
X-RAY DIFFRACTIONf_dihedral_angle_d25.55946491
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.39-3.46100.57674X-RAY DIFFRACTION0.1
3.51-3.55100.60045X-RAY DIFFRACTION0.27
3.56-3.640.376820.33797X-RAY DIFFRACTION0.28
3.66-3.750.439370.399136X-RAY DIFFRACTION3.94
3.75-3.870.3948880.35231395X-RAY DIFFRACTION37.41
3.87-4.010.3971880.32043204X-RAY DIFFRACTION84.84
4.01-4.170.39232160.31923712X-RAY DIFFRACTION97.81
4.17-4.360.37181970.32643783X-RAY DIFFRACTION99.38
4.36-4.590.32161910.31433821X-RAY DIFFRACTION99.78
4.59-4.870.33762030.30713804X-RAY DIFFRACTION99.73
4.87-5.240.35621820.30823853X-RAY DIFFRACTION99.88
5.24-5.760.32041880.3323861X-RAY DIFFRACTION99.88
5.76-6.570.35092140.33953833X-RAY DIFFRACTION99.8
6.57-8.20.30182080.33183879X-RAY DIFFRACTION99.98
8.2-21.410.25872120.25753928X-RAY DIFFRACTION98.03

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