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- PDB-6dcc: Structure of methylphosphate capping enzyme methyltransferase dom... -

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Basic information

Entry
Database: PDB / ID: 6dcc
TitleStructure of methylphosphate capping enzyme methyltransferase domain in complex with 5' end of 7SK RNA
Components
  • 7SK snRNA methylphosphate capping enzyme
  • human 7SK RNA stem-loop 1 proximal methylated
KeywordsTransferase/RNA / RNA methyl transferase / MePCE / 7SK RNA / Transferase-RNA complex
Function / homology
Function and homology information


RNA 5'-gamma-phosphate methyltransferase activity / snRNA metabolic process / snRNA modification / 7SK snRNP / positive regulation of snRNA transcription by RNA polymerase II / 7SK snRNA binding / snRNA binding / RNA methyltransferase activity / positive regulation of protein localization to Cajal body / RNA methylation ...RNA 5'-gamma-phosphate methyltransferase activity / snRNA metabolic process / snRNA modification / 7SK snRNP / positive regulation of snRNA transcription by RNA polymerase II / 7SK snRNA binding / snRNA binding / RNA methyltransferase activity / positive regulation of protein localization to Cajal body / RNA methylation / O-methyltransferase activity / S-adenosylmethionine-dependent methyltransferase activity / positive regulation of G1/S transition of mitotic cell cycle / Transferases; Transferring one-carbon groups; Methyltransferases / ribonucleoprotein complex / negative regulation of transcription by RNA polymerase II / RNA binding / nucleus
Similarity search - Function
RNA methyltransferase bin3, C-terminal / Bin3-type S-adenosyl-L-methionine binding domain / RNA methyltransferase Bin3-like / Bicoid-interacting protein 3 (Bin3) / Bin3-type S-adenosyl-L-methionine (SAM) domain profile. / Methyltransferase domain / Vaccinia Virus protein VP39 / S-adenosyl-L-methionine-dependent methyltransferase superfamily / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
S-ADENOSYL-L-HOMOCYSTEINE / RNA / RNA (> 10) / 7SK snRNA methylphosphate capping enzyme
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 2.1 Å
Model detailsTelomerase protein in complex with RNA
AuthorsYang, Y. / Eichhorn, C. / Cascio, D. / Feigon, J.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research Institute (NIH/NHGRI)GM107567 United States
CitationJournal: Nat. Chem. Biol. / Year: 2019
Title: Structural basis of 7SK RNA 5'-gamma-phosphate methylation and retention by MePCE.
Authors: Yang, Y. / Eichhorn, C.D. / Wang, Y. / Cascio, D. / Feigon, J.
History
DepositionMay 4, 2018Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 19, 2018Provider: repository / Type: Initial release
Revision 1.1Jan 2, 2019Group: Data collection / Database references / Category: citation / citation_author
Item: _citation.journal_abbrev / _citation.pdbx_database_id_PubMed ..._citation.journal_abbrev / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.2Jan 30, 2019Group: Data collection / Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year / _citation_author.identifier_ORCID
Revision 1.3Dec 18, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.4Oct 11, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 7SK snRNA methylphosphate capping enzyme
B: human 7SK RNA stem-loop 1 proximal methylated
hetero molecules


Theoretical massNumber of molelcules
Total (without water)47,3274
Polymers46,8472
Non-polymers4802
Water1,63991
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: native gel electrophoresis
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area2020 Å2
ΔGint-21 kcal/mol
Surface area15970 Å2
2
A: 7SK snRNA methylphosphate capping enzyme
B: human 7SK RNA stem-loop 1 proximal methylated
hetero molecules

A: 7SK snRNA methylphosphate capping enzyme
B: human 7SK RNA stem-loop 1 proximal methylated
hetero molecules

A: 7SK snRNA methylphosphate capping enzyme
B: human 7SK RNA stem-loop 1 proximal methylated
hetero molecules


Theoretical massNumber of molelcules
Total (without water)141,98112
Polymers140,5406
Non-polymers1,4416
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation2_565-y,x-y+1,z1
crystal symmetry operation3_455-x+y-1,-x,z1
Buried area11150 Å2
ΔGint-102 kcal/mol
Surface area42820 Å2
MethodPISA
Unit cell
Length a, b, c (Å)119.690, 119.690, 77.820
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number173
Space group name H-MP63

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Components

#1: Protein 7SK snRNA methylphosphate capping enzyme / MePCE / Bicoid-interacting protein 3 homolog / Bin3 homolog


Mass: 35122.797 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MEPCE, BCDIN3 / Plasmid: pET28a / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3)
References: UniProt: Q7L2J0, Transferases; Transferring one-carbon groups; Methyltransferases
#2: RNA chain human 7SK RNA stem-loop 1 proximal methylated


Mass: 11723.851 Da / Num. of mol.: 1 / Source method: obtained synthetically
Details: in vitro T7 polymerase transciption from synthetic DNA template, 5' methylated by cocrytallization with MePCE and SAM.
Source: (synth.) Homo sapiens (human)
#3: Chemical ChemComp-SAH / S-ADENOSYL-L-HOMOCYSTEINE / S-Adenosyl-L-homocysteine


Type: L-peptide linking / Mass: 384.411 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C14H20N6O5S
#4: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: SO4
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 91 / 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: 3.61 Å3/Da / Density % sol: 65.9 %
Crystal growTemperature: 277 K / Method: vapor diffusion, hanging drop / pH: 4.2
Details: crystallization solution 0.2M Lithium sulfate, 0.1M phosphate/citrate, pH 4.2, 25% w/v PEG mixed 1:2 volume ratio to 10 mg/mL protein:RNA complex. Crystals were soaked in crystallization ...Details: crystallization solution 0.2M Lithium sulfate, 0.1M phosphate/citrate, pH 4.2, 25% w/v PEG mixed 1:2 volume ratio to 10 mg/mL protein:RNA complex. Crystals were soaked in crystallization solution supplemented with 1.2mM MgCl2 for 2hrs prior to freezing.

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-C / Wavelength: 0.9792 Å
DetectorType: DECTRIS PILATUS 6M-F / Detector: PIXEL / Date: Mar 22, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9792 Å / Relative weight: 1
ReflectionResolution: 2.1→51.827 Å / Num. obs: 37061 / % possible obs: 99.8 % / Redundancy: 5.132 % / Biso Wilson estimate: 44.57 Å2 / CC1/2: 0.999 / Rmerge(I) obs: 0.056 / Rrim(I) all: 0.062 / Χ2: 1.05 / Net I/σ(I): 15.61 / Num. measured all: 190193 / Scaling rejects: 9
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsMean I/σ(I) obsNum. measured obsNum. possibleNum. unique obsCC1/2Rrim(I) all% possible all
2.1-2.155.1960.781.914091271727120.7110.8799.8
2.15-2.215.1440.6642.2613651265526540.740.74100
2.21-2.284.9660.5472.7312818258125810.7940.612100
2.28-2.354.9160.3663.7712398252525220.9340.41199.9
2.35-2.425.3610.3114.8313102244524440.9450.345100
2.42-2.515.3040.2456.0212545236523650.9630.272100
2.51-2.65.2180.2047.3111866227622740.970.22799.9
2.6-2.715.1580.1559.3111213217621740.9820.17399.9
2.71-2.834.780.1211.3810058211121040.9880.13599.7
2.83-2.975.3190.09515.410734202220180.9930.10599.8
2.97-3.135.3510.07619.2610269192019190.9960.08499.9
3.13-3.325.2280.05625.49599183818360.9970.06299.9
3.32-3.555.0560.04529.428555169216920.9980.05100
3.55-3.834.8470.03932.237678158815840.9980.04399.7
3.83-4.25.3470.03438.27817146614620.9990.03799.7
4.2-4.75.2060.03240.056872132213200.9990.03599.8
4.7-5.424.780.03239.045660118911840.9980.03699.6
5.42-6.645.1690.03439.135190100510040.9990.03799.9
6.64-9.395.1660.02844.1139787777700.9990.03199.1
9.39-51.8274.7490.02746.2420994474420.9980.03198.9

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Phasing

PhasingMethod: molecular replacement
Phasing MRModel details: Phaser MODE: MR_AUTO
Highest resolutionLowest resolution
Rotation6.41 Å61.97 Å
Translation6.41 Å61.97 Å

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Processing

Software
NameVersionClassification
PHENIXrefinement
XSCALEdata scaling
PHASER2.7.17phasing
PDB_EXTRACT3.24data extraction
XDSdata reduction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5UNA

5una


Resolution: 2.1→51.827 Å / SU ML: 0.23 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 21.04 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.1988 3706 10 %
Rwork0.1748 33346 -
obs0.1772 37052 99.83 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 185.38 Å2 / Biso mean: 54.1953 Å2 / Biso min: 27.21 Å2
Refinement stepCycle: final / Resolution: 2.1→51.827 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1759 776 31 91 2657
Biso mean--37.82 47.73 -
Num. residues----257
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0072716
X-RAY DIFFRACTIONf_angle_d0.9943871
X-RAY DIFFRACTIONf_chiral_restr0.048449
X-RAY DIFFRACTIONf_plane_restr0.005351
X-RAY DIFFRACTIONf_dihedral_angle_d14.0771500
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 26

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
2.1-2.12760.28161400.245712591399100
2.1276-2.15680.24041410.220512701411100
2.1568-2.18760.25871420.230712741416100
2.1876-2.22020.25021420.228612791421100
2.2202-2.25490.26241420.236312761418100
2.2549-2.29190.23751410.196112721413100
2.2919-2.33140.23841440.210312891433100
2.3314-2.37380.23551400.206212651405100
2.3738-2.41950.2551440.194112941438100
2.4195-2.46890.23481410.199812701411100
2.4689-2.52250.23091420.194212791421100
2.5225-2.58120.24071420.201412751417100
2.5812-2.64580.22551440.197812951439100
2.6458-2.71730.21921410.209712721413100
2.7173-2.79720.26191430.207212811424100
2.7972-2.88750.26771420.214912821424100
2.8875-2.99070.26621430.219212821425100
2.9907-3.11040.23491420.215812831425100
3.1104-3.2520.26831430.211512841427100
3.252-3.42340.23741420.185412831425100
3.4234-3.63780.171440.165112941438100
3.6378-3.91860.13831430.147412841427100
3.9186-4.31280.1481420.129812831425100
4.3128-4.93650.14391450.124512991444100
4.9365-6.21780.18521440.159112951439100
6.2178-51.84310.17061470.16061327147499
Refinement TLS params.Method: refined / Origin x: -50.2292 Å / Origin y: 14.1813 Å / Origin z: -3.2789 Å
111213212223313233
T0.3318 Å20.0175 Å20.0213 Å2-0.2543 Å2-0.0028 Å2--0.2893 Å2
L1.2142 °20.6497 °2-0.1027 °2-1.896 °2-0.5418 °2--0.7424 °2
S-0.0465 Å °0.0325 Å °-0.2102 Å °-0.0022 Å °-0.0521 Å °-0.242 Å °0.1078 Å °0.0986 Å °0.1219 Å °
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1allA412 - 687
2X-RAY DIFFRACTION1allB1 - 116
3X-RAY DIFFRACTION1allC1
4X-RAY DIFFRACTION1allD1
5X-RAY DIFFRACTION1allF1 - 91

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