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- PDB-3nh2: Crystal structure of RNase T in complex with a stem DNA with a 3'... -

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Basic information

Entry
Database: PDB / ID: 3nh2
TitleCrystal structure of RNase T in complex with a stem DNA with a 3' overhang
Components
  • 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'
  • Ribonuclease T
KeywordsHYDROLASE/DNA / exoribonuclease / RNA processing / RNA maturation / protein-DNA interactions / protein-DNA complex / exo-nuclease / HYDROLASE-DNA complex
Function / homology
Function and homology information


rRNA 3'-end processing / tRNA 3'-end processing / regulatory ncRNA 3'-end processing / single-stranded DNA 3'-5' DNA exonuclease activity / DNA replication proofreading / 3'-5' exonuclease activity / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / 3'-5'-RNA exonuclease activity / nucleic acid binding / DNA damage response ...rRNA 3'-end processing / tRNA 3'-end processing / regulatory ncRNA 3'-end processing / single-stranded DNA 3'-5' DNA exonuclease activity / DNA replication proofreading / 3'-5' exonuclease activity / Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters / 3'-5'-RNA exonuclease activity / nucleic acid binding / DNA damage response / magnesium ion binding / protein homodimerization activity / identical protein binding / cytosol
Similarity search - Function
Ribonuclease T / Exonuclease / Exonuclease, RNase T/DNA polymerase III / EXOIII / Ribonuclease H-like superfamily/Ribonuclease H / Nucleotidyltransferase; domain 5 / Ribonuclease H superfamily / Ribonuclease H-like superfamily / 2-Layer Sandwich / Alpha Beta
Similarity search - Domain/homology
DNA / Ribonuclease T
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
synthetic construct (others)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.3 Å
AuthorsHsiao, Y.-Y. / Yuan, H.S.
CitationJournal: Nat.Chem.Biol. / Year: 2011
Title: Structural basis for RNA trimming by RNase T in stable RNA 3'-end maturation
Authors: Hsiao, Y.-Y. / Yang, C.-C. / Lin, C.L. / Lin, J.L.J. / Duh, Y. / Yuan, H.S.
History
DepositionJun 14, 2010Deposition site: RCSB / Processing site: PDBJ
Revision 1.0Feb 16, 2011Provider: repository / Type: Initial release
Revision 1.1Jul 13, 2011Group: Version format compliance
Revision 1.2Dec 21, 2011Group: Database references
Revision 1.3May 31, 2023Group: Database references / Derived calculations / Source and taxonomy
Category: database_2 / pdbx_entity_src_syn ...database_2 / pdbx_entity_src_syn / pdbx_struct_assembly / pdbx_struct_assembly_gen / pdbx_struct_assembly_prop / struct_ref_seq_dif
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_ref_seq_dif.details
Revision 1.4Nov 8, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Ribonuclease T
B: Ribonuclease T
C: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'
D: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'
E: Ribonuclease T
F: Ribonuclease T
G: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'
H: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'


Theoretical massNumber of molelcules
Total (without water)111,2028
Polymers111,2028
Non-polymers00
Water6,215345
1
A: Ribonuclease T
B: Ribonuclease T
C: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'
D: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'


Theoretical massNumber of molelcules
Total (without water)55,6014
Polymers55,6014
Non-polymers00
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
E: Ribonuclease T
F: Ribonuclease T
G: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'
H: 5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'


Theoretical massNumber of molelcules
Total (without water)55,6014
Polymers55,6014
Non-polymers00
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)60.379, 62.560, 62.657
Angle α, β, γ (deg.)82.77, 82.92, 66.13
Int Tables number1
Space group name H-MP1

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Components

#1: Protein
Ribonuclease T / RNase T / Exoribonuclease T


Mass: 25719.035 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Strain: JM109 / ATCC 53323 / Gene: rnt / Plasmid: pET28a / Production host: Escherichia coli (E. coli) / Strain (production host): RIPL
References: UniProt: P30014, Hydrolases; Acting on ester bonds; Exoribonucleases producing 5'-phosphomonoesters
#2: DNA chain
5'-D(P*TP*TP*AP*CP*AP*AP*C)-3'


Mass: 2081.412 Da / Num. of mol.: 4 / Source method: obtained synthetically / Details: ssDNA / Source: (synth.) synthetic construct (others)
#3: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 345 / 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: 1.92 Å3/Da / Density % sol: 36.09 %
Crystal growTemperature: 298 K / Method: vapor diffusion, hanging drop / pH: 5.5
Details: 0.1% w/v n-Octyl-beta-D-glucoside, 0.1M Sodium citrate tribasic dihydrate, pH 5.5, 22% PEG 4000, VAPOR DIFFUSION, HANGING DROP, temperature 298K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: NSRRC / Beamline: BL13B1 / Wavelength: 0.999 Å
DetectorType: ADSC QUANTUM 315 / Detector: CCD / Date: Nov 14, 2009
RadiationMonochromator: SAGITALLY FOCUSED Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.999 Å / Relative weight: 1
ReflectionResolution: 2.3→30 Å / Num. all: 35579 / Num. obs: 35579 / % possible obs: 96.5 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 2.6 % / Rsym value: 0.1 / Net I/σ(I): 9.7
Reflection shellResolution: 2.3→2.38 Å / Redundancy: 2.5 % / Mean I/σ(I) obs: 2.4 / Num. unique all: 3544 / Rsym value: 0.41 / % possible all: 96.5

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Processing

Software
NameVersionClassification
HKL-2000data collection
AMoREphasing
PHENIX(phenix.refine: 1.6_289)refinement
HKL-2000data reduction
HKL-2000data scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 3NGY
Resolution: 2.3→26.761 Å / Occupancy max: 1 / Occupancy min: 1 / FOM work R set: 0.806 / SU ML: 0.34 / Cross valid method: THROUGHOUT / σ(F): 1.97 / Phase error: 26.72 / Stereochemistry target values: ML
RfactorNum. reflection% reflectionSelection details
Rfree0.2409 2693 7.6 %RANDOM
Rwork0.1858 32763 --
all0.2085 35456 --
obs0.19 35456 96.32 %-
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL / Bsol: 40.97 Å2 / ksol: 0.4 e/Å3
Displacement parametersBiso max: 78.83 Å2 / Biso mean: 26.2313 Å2 / Biso min: 8.14 Å2
Baniso -1Baniso -2Baniso -3
1-1.157 Å2-0.5541 Å2-0.5855 Å2
2---1.0876 Å22.4737 Å2
3----0.0694 Å2
Refinement stepCycle: LAST / Resolution: 2.3→26.761 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6288 524 0 345 7157
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0037023
X-RAY DIFFRACTIONf_angle_d0.7369618
X-RAY DIFFRACTIONf_dihedral_angle_d17.4352514
X-RAY DIFFRACTIONf_chiral_restr0.051064
X-RAY DIFFRACTIONf_plane_restr0.0021176
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.3-2.34180.26581390.19831697X-RAY DIFFRACTION95
2.3418-2.38680.33081460.20591733X-RAY DIFFRACTION97
2.3868-2.43550.2641400.20831723X-RAY DIFFRACTION97
2.4355-2.48840.28511390.20251759X-RAY DIFFRACTION96
2.4884-2.54630.34341370.21281718X-RAY DIFFRACTION96
2.5463-2.60990.27441540.20531761X-RAY DIFFRACTION97
2.6099-2.68040.26481180.2021704X-RAY DIFFRACTION97
2.6804-2.75920.2881430.20391751X-RAY DIFFRACTION97
2.7592-2.84820.29881420.2051755X-RAY DIFFRACTION97
2.8482-2.94980.27621350.19221731X-RAY DIFFRACTION97
2.9498-3.06780.24621490.18581716X-RAY DIFFRACTION96
3.0678-3.20720.23781380.18011735X-RAY DIFFRACTION97
3.2072-3.3760.22691470.17141750X-RAY DIFFRACTION97
3.376-3.5870.20521470.16561714X-RAY DIFFRACTION97
3.587-3.86320.21121330.15541723X-RAY DIFFRACTION95
3.8632-4.25060.19391470.16621710X-RAY DIFFRACTION96
4.2506-4.86250.19181540.14551732X-RAY DIFFRACTION96
4.8625-6.11420.18321360.17451674X-RAY DIFFRACTION94
6.1142-26.76230.22851490.19631677X-RAY DIFFRACTION95

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