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- PDB-5lel: Crystal structure of DARPin-DARPin rigid fusion, variant DD_Off7_... -

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Basic information

Entry
Database: PDB / ID: 5lel
TitleCrystal structure of DARPin-DARPin rigid fusion, variant DD_Off7_10_3G124 in complex with Maltose-binding Protein and Green Fluorescent Protein
Components
  • DD_Off7_10_3G124
  • Green fluorescent protein
  • Maltose-binding periplasmic protein
KeywordsFLUORESCENT PROTEIN / X-ray crystallography / designed ankyrin repeat proteins / protein design / protein engineering / rigid domain fusions
Function / homology
Function and homology information


carbohydrate transmembrane transporter activity / bioluminescence / generation of precursor metabolites and energy / outer membrane-bounded periplasmic space
Similarity search - Function
Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / Green Fluorescent Protein / Green fluorescent protein / Bacterial extracellular solute-binding protein / Green fluorescent protein, GFP / Bacterial extracellular solute-binding protein / Green fluorescent protein-related / Green fluorescent protein ...Maltose/Cyclodextrin ABC transporter, substrate-binding protein / Solute-binding family 1, conserved site / Bacterial extracellular solute-binding proteins, family 1 signature. / Green Fluorescent Protein / Green fluorescent protein / Bacterial extracellular solute-binding protein / Green fluorescent protein, GFP / Bacterial extracellular solute-binding protein / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Beta Barrel / Mainly Beta
Similarity search - Domain/homology
Maltose/maltodextrin-binding periplasmic protein / Green fluorescent protein
Similarity search - Component
Biological speciessynthetic construct (others)
Escherichia coli K-12 (bacteria)
Aequorea victoria (jellyfish)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.1 Å
AuthorsBatyuk, A. / Wu, Y. / Mittl, P.R. / Plueckthun, A.
Funding support Switzerland, 2items
OrganizationGrant numberCountry
Swiss National Science Foundation310030B_166676 Switzerland
European Research CouncilNEXTBINDERS
CitationJournal: Sci Rep / Year: 2017
Title: Rigidly connected multispecific artificial binders with adjustable geometries.
Authors: Wu, Y. / Batyuk, A. / Honegger, A. / Brandl, F. / Mittl, P.R.E. / Pluckthun, A.
History
DepositionJun 30, 2016Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 15, 2017Provider: repository / Type: Initial release
Revision 1.1Jan 24, 2018Group: Source and taxonomy / Category: entity_src_gen
Item: _entity_src_gen.pdbx_gene_src_ncbi_taxonomy_id / _entity_src_gen.pdbx_gene_src_scientific_name / _entity_src_gen.pdbx_host_org_ncbi_taxonomy_id
Revision 1.2Mar 20, 2019Group: Data collection / Database references / Category: citation / citation_author / pdbx_database_proc
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name
Revision 1.3Oct 16, 2019Group: Data collection / Category: reflns_shell

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DD_Off7_10_3G124
B: Maltose-binding periplasmic protein
C: Green fluorescent protein
D: DD_Off7_10_3G124
E: Maltose-binding periplasmic protein
F: Green fluorescent protein
G: DD_Off7_10_3G124
H: Maltose-binding periplasmic protein
I: Green fluorescent protein


Theoretical massNumber of molelcules
Total (without water)319,0319
Polymers319,0319
Non-polymers00
Water0
1
A: DD_Off7_10_3G124
B: Maltose-binding periplasmic protein
C: Green fluorescent protein


Theoretical massNumber of molelcules
Total (without water)106,3443
Polymers106,3443
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
D: DD_Off7_10_3G124
E: Maltose-binding periplasmic protein
F: Green fluorescent protein


Theoretical massNumber of molelcules
Total (without water)106,3443
Polymers106,3443
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
G: DD_Off7_10_3G124
H: Maltose-binding periplasmic protein
I: Green fluorescent protein


Theoretical massNumber of molelcules
Total (without water)106,3443
Polymers106,3443
Non-polymers00
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)235.000, 103.140, 156.840
Angle α, β, γ (deg.)90.00, 131.35, 90.00
Int Tables number5
Space group name H-MC121

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Components

#1: Protein DD_Off7_10_3G124


Mass: 34951.281 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli) / Strain (production host): K-12 / Variant (production host): XL1-Blue
#2: Protein Maltose-binding periplasmic protein / MBP / MMBP / Maltodextrin-binding protein


Mass: 43172.637 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli K-12 (bacteria) / Gene: malE, Z5632, ECs5017 / Production host: Escherichia coli (E. coli) / Strain (production host): K-12 / Variant (production host): XL1-Blue / References: UniProt: P0AEY0
#3: Protein Green fluorescent protein /


Mass: 28219.721 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Aequorea victoria (jellyfish) / Gene: GFP / Production host: Escherichia coli (E. coli) / Strain (production host): K-12 / Variant (production host): XL1-Blue / References: UniProt: P42212

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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: 2.24 Å3/Da / Density % sol: 45 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 8
Details: PEG3350 23.2% w/v, Sodium acetate 0.2 M, Bis-Tris propane 0.1 M, pH 8.0

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X06SA / Wavelength: 1 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Mar 25, 2013
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 3.1→46.63 Å / Num. obs: 51171 / % possible obs: 99.7 % / Redundancy: 6.7 % / CC1/2: 0.993 / Net I/σ(I): 5.81

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Processing

Software
NameVersionClassification
PHENIX(DEV_2429)refinement
XDSdata reduction
XSCALEdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 3.1→46.63 Å / SU ML: 0.64 / Cross valid method: FREE R-VALUE / σ(F): 1.99 / Phase error: 36.86
RfactorNum. reflection% reflection
Rfree0.306 2557 5 %
Rwork0.258 --
obs0.261 51133 99.7 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Refinement stepCycle: LAST / Resolution: 3.1→46.63 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms21046 0 0 0 21046
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00221514
X-RAY DIFFRACTIONf_angle_d0.49929197
X-RAY DIFFRACTIONf_dihedral_angle_d8.68412731
X-RAY DIFFRACTIONf_chiral_restr0.043223
X-RAY DIFFRACTIONf_plane_restr0.0033821
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.0999-3.15960.38891410.40332671X-RAY DIFFRACTION99
3.1596-3.2240.39071410.39482675X-RAY DIFFRACTION99
3.224-3.29410.40431410.37532677X-RAY DIFFRACTION99
3.2941-3.37070.40291390.37312649X-RAY DIFFRACTION100
3.3707-3.4550.43671420.36242703X-RAY DIFFRACTION100
3.455-3.54840.35721420.34412696X-RAY DIFFRACTION100
3.5484-3.65280.42371410.32812670X-RAY DIFFRACTION100
3.6528-3.77060.33871410.31042680X-RAY DIFFRACTION100
3.7706-3.90530.3641420.30332693X-RAY DIFFRACTION100
3.9053-4.06160.35591420.28452705X-RAY DIFFRACTION100
4.0616-4.24630.30521410.26682689X-RAY DIFFRACTION100
4.2463-4.470.31021430.23552713X-RAY DIFFRACTION100
4.47-4.74980.26391420.21392710X-RAY DIFFRACTION100
4.7498-5.11620.26931420.22442692X-RAY DIFFRACTION100
5.1162-5.63020.32981420.24062706X-RAY DIFFRACTION100
5.6302-6.44310.2971450.25142742X-RAY DIFFRACTION100
6.4431-8.11070.29071430.21342720X-RAY DIFFRACTION100
8.1107-46.63530.21181470.18382785X-RAY DIFFRACTION100
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL112)L122)L132)L222)L232)L332)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T112)T122)T132)T222)T232)T332)Origin x (Å)Origin y (Å)Origin z (Å)
10.67510.77980.25711.6219-0.04610.2803-0.8628-0.7177-0.0299-0.58010.7166-0.01820.12471.30470.05030.22410.3274-0.05291.384-0.47130.959472.1711-37.609822.8368
20.88130.0223-0.29240.7883-0.47220.1914-0.4507-0.4151-0.60180.03840.60870.23140.04-0.40430.07560.478-0.1276-0.11510.8513-0.3780.551549.5233-31.48815.955
30.0887-0.39740.15070.934-0.74760.61490.3435-0.20930.06680.23260.07740.4252-0.21440.3653-00.5255-0.02820.00290.72510.03780.58325.0238-35.731339.2592
40.03590.1259-0.06140.1595-0.1450.0699-0.69770.2829-0.0896-0.1098-0.466-0.7307-0.42710.0256-01.5503-0.2298-0.03031.87120.07951.028199.5702-23.9261-30.3226
50.68320.3195-0.91180.6838-0.59971.175-0.91180.4066-0.0496-0.19510.04780.33030.4979-0.6271-0.45320.82040.5567-0.6055-0.04220.5840.733384.7026-16.5902-25.0262
60.2909-0.16530.22570.50490.81671.8623-0.4865-0.91080.053-0.2773-0.8095-0.25960.3876-0.6057-0.59520.27780.95850.0636-1.054-0.4990.96670.9267-35.3974-2.8086
70.6434-0.0676-1.2581.6156-1.12731.80380.0173-0.33330.2328-0.5157-0.29860.0109-0.13130.35-0.04730.82580.0466-0.13660.5339-0.00580.777977.664-24.7783-13.0455
80.3280.6848-0.08930.96290.15290.38960.0225-0.59780.0367-0.49050.4780.1341-0.16960.74780.0610.51840.01360.12690.6798-0.06490.579249.1833-41.773244.3126
90.1032-0.29-0.010.79890.00060.0424-0.57120.13790.84311.61890.01080.7498-1.2945-0.1256-0.02090.89490.0459-0.05790.5471-0.17630.799639.3418-32.127859.0904
100.2039-0.1874-0.03130.0629-0.04780.5427-0.14860.19160.0897-0.30520.3994-0.6040.63710.7324-0.51730.0496-0.60910.070.7044-0.35590.950852.198-41.778756.4617
110.40.0289-0.34720.05280.05540.33630.1959-0.12980.2546-0.468-0.0210.10360.12211.2427-0.00150.528-0.04390.04170.9546-0.19450.833956.7453-40.222348.8965
121.5271-0.7634-1.15162.0395-0.12040.6825-0.2934-1.3116-0.23960.1770.3850.87640.30120.92980.30680.4748-0.00450.01240.6873-0.20560.351948.739-49.071257.0001
130.26830.1697-0.02290.060.04750.0514-0.5365-0.2552-0.0104-0.2085-0.19310.176-0.12220.4816-0.0140.5481-0.0210.02890.5524-0.21620.40140.5532-44.849945.9761
140.504-0.0516-0.46990.15020.06750.60780.46910.97360.02970.3802-0.24040.7612-0.48130.2740.10540.6278-0.030.11260.35260.23780.787612.3708-4.22259.174
150.5852-0.01910.448-0.08440.10110.713-0.1140.01030.26350.08740.13050.05130.5868-0.0141-0.00010.9086-0.10540.23610.5670.0730.72731.4793-24.867731.9573
160.36820.3648-0.0770.47380.4020.51970.20990.12040.00180.2216-0.46460.026-0.0013-0.1884-00.68120.06440.10180.77810.06710.4254-10.0758-14.95352.3246
170.11230.1220.11310.19030.12060.10220.1226-0.5660.64280.15910.06870.25560.29460.06180.00041.24550.22970.23841.02290.2110.788946.5264-14.9492-41.2708
180.84750.4928-0.50810.74980.0740.51440.25840.1183-0.0845-0.15-0.2636-0.48310.55650.1112-01.06960.24070.04080.68020.09740.520735.1091-24.8078-33.0171
190.27870.1389-0.20210.46240.08170.11050.451-0.1610.2446-0.0486-0.01850.39190.46130.46330.00020.66360.013-0.06360.4833-0.00810.68529.3501-12.7881-3.8216
200.4884-0.5835-0.06961.388-0.55061.46890.36770.00730.0728-0.16350.0235-0.0330.12820.1035-0.00010.5980.0099-0.05980.2939-0.03570.570528.6144-17.1517-15.0744
210.3661-0.51630.06710.5218-0.31090.47560.1262-0.0521-0.05750.2744-0.4462-0.24850.73620.2375-0.00030.88590.22730.1110.86450.04510.687245.2382-22.2715-25.5041
220.42910.4444-0.07730.482-0.22080.2698-0.2591.02570.2511-1.12360.01030.16640.44450.7022-0.35481.50190.6064-0.34941.0742-0.26380.042616.1901-17.2398-20.835
230.0207-0.0151-0.03540.00370.0150.07050.28260.16410.19430.2638-0.49590.2411-0.7179-0.14870.00191.0920.1836-0.08811.0914-0.3190.59538.9504-21.6533-14.4282
240.23460.14020.42670.68460.63511.36590.49490.15680.1528-0.1892-0.0764-0.5065-0.0973-1.28240.12730.71840.0210.19130.76920.29110.7785-5.7730.608132.082
250.3033-0.44220.24080.6814-0.03330.58010.11610.4554-0.06320.5150.1919-0.0301-0.2245-0.260.00010.56970.15010.09690.5341-0.04080.5538-9.05064.305638.1858
260.8524-0.4332-0.38511.3761.32690.99720.69370.33990.7363-0.2327-0.15210.0881-0.2567-0.21830.01090.62670.05320.13220.74160.21790.6978-8.981412.263437.6207
270.34030.63910.38562.00271.20330.5804-0.04610.55520.3709-0.06980.5233-0.9563-0.0034-0.36770.51760.47460.045-0.05670.65590.33680.3011-4.2594-0.030342.4402
280.41690.5305-0.17960.65670.2480.7223-0.21220.10810.1634-0.4486-0.3288-0.1060.55360.6608-00.93150.1662-0.08150.70820.22530.643371.379-12.715534.7957
290.9106-0.4315-1.15111.3892-0.20211.2705-0.40250.16070.1250.05020.3368-0.94060.0342-0.3427-0.19280.69940.1123-0.14630.63080.11611.201886.86985.73533.02
300.12890.0501-0.10950.0220.0040.07181.2373-0.69310.40960.48950.2762-0.1054-0.26190.05310.00191.73470.61450.27072.39850.36371.622923.5143-0.6872.5004
312.01050.33890.77020.47750.47190.54721.07870.8754-0.11781.8363-0.77730.27380.0004-0.18320.40351.74660.72160.58652.27710.96061.403419.8745-12.677966.0584
320.088-0.07730.1470.3437-0.02580.4382-1.1722-1.9445-0.84560.68910.69220.5642-0.0053-0.1721-0.02690.68950.24390.08481.07950.04151.097524.47290.352453.3671
330.7037-0.3023-0.8133-0.13530.39250.8194-0.2069-0.89940.27340.11140.09670.05730.03550.000400.72620.1089-0.16350.68720.0870.813748.1508-3.348242.8001
342.50620.9662-0.20050.59450.14680.3628-0.4934-3.0330.6524-0.1177-0.39080.34390.20090.0581-0.40950.73250.7474-0.54251.83090.17440.232941.4173-0.256754.6593
350.35420.07960.04170.6182-0.4440.3096-0.50760.1930.06950.02490.7794-0.42140.28230.2255-0.00631.03060.42540.01971.3197-0.44431.160230.28966.73360.2922
360.07970.10140.07930.10080.09480.0906-0.3602-0.5936-1.0914-0.58870.0490.4348-0.259-0.5872-0.00420.59890.0450.15660.85480.04480.912439.2795-14.984435.8246
371.84510.9467-0.82220.579-0.4160.3211-1.32760.05450.27030.87680.9214-0.15530.9444-0.1869-0.28721.31180.3327-0.24620.74280.02250.547993.703-11.743613.8365
380.2655-0.0164-0.10280.00360.01430.14860.0021-0.5373-0.10170.13480.0847-0.06050.43730.37130.00622.31841.755-1.18971.7913-0.38691.14299.9907-9.092922.9299
391.21591.1086-1.09131.1566-1.00791.0482-1.5533-0.6999-0.22070.16050.4904-0.26281.06660.2089-0.81931.22810.5578-0.65631.12930.0231.4437102.8459-15.491615.8348
400.72990.4592-0.55810.5372-0.43660.5818-0.9466-1.3067-0.19870.59840.7648-0.7417-0.1674-0.8983-0.14441.21340.09-0.53081.2426-0.19771.8257112.1581-15.743613.8108
410.1441-0.0418-0.3181-0.1391-0.09630.5466-0.8909-0.3667-0.297-0.39660.7266-0.3605-0.33030.2221-0.01060.99230.1083-0.18171.0718-0.29150.9662103.4568-11.601611.878
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection details
1X-RAY DIFFRACTION1CHAIN 'A' AND (RESID 12 THROUGH 101 )
2X-RAY DIFFRACTION2CHAIN 'A' AND (RESID 102 THROUGH 180 )
3X-RAY DIFFRACTION3CHAIN 'A' AND (RESID 181 THROUGH 324 )
4X-RAY DIFFRACTION4CHAIN 'B' AND (RESID 5 THROUGH 42 )
5X-RAY DIFFRACTION5CHAIN 'B' AND (RESID 43 THROUGH 118 )
6X-RAY DIFFRACTION6CHAIN 'B' AND (RESID 119 THROUGH 164 )
7X-RAY DIFFRACTION7CHAIN 'B' AND (RESID 165 THROUGH 372 )
8X-RAY DIFFRACTION8CHAIN 'C' AND (RESID 2 THROUGH 68 )
9X-RAY DIFFRACTION9CHAIN 'C' AND (RESID 69 THROUGH 81 )
10X-RAY DIFFRACTION10CHAIN 'C' AND (RESID 82 THROUGH 104 )
11X-RAY DIFFRACTION11CHAIN 'C' AND (RESID 105 THROUGH 128 )
12X-RAY DIFFRACTION12CHAIN 'C' AND (RESID 129 THROUGH 198 )
13X-RAY DIFFRACTION13CHAIN 'C' AND (RESID 199 THROUGH 230 )
14X-RAY DIFFRACTION14CHAIN 'D' AND (RESID 12 THROUGH 158 )
15X-RAY DIFFRACTION15CHAIN 'D' AND (RESID 159 THROUGH 225 )
16X-RAY DIFFRACTION16CHAIN 'D' AND (RESID 226 THROUGH 324 )
17X-RAY DIFFRACTION17CHAIN 'E' AND (RESID 5 THROUGH 42 )
18X-RAY DIFFRACTION18CHAIN 'E' AND (RESID 43 THROUGH 118 )
19X-RAY DIFFRACTION19CHAIN 'E' AND (RESID 119 THROUGH 164 )
20X-RAY DIFFRACTION20CHAIN 'E' AND (RESID 165 THROUGH 273 )
21X-RAY DIFFRACTION21CHAIN 'E' AND (RESID 274 THROUGH 333 )
22X-RAY DIFFRACTION22CHAIN 'E' AND (RESID 334 THROUGH 351 )
23X-RAY DIFFRACTION23CHAIN 'E' AND (RESID 352 THROUGH 373 )
24X-RAY DIFFRACTION24CHAIN 'F' AND (RESID 2 THROUGH 48 )
25X-RAY DIFFRACTION25CHAIN 'F' AND (RESID 49 THROUGH 81 )
26X-RAY DIFFRACTION26CHAIN 'F' AND (RESID 82 THROUGH 198 )
27X-RAY DIFFRACTION27CHAIN 'F' AND (RESID 199 THROUGH 230 )
28X-RAY DIFFRACTION28CHAIN 'G' AND (RESID 13 THROUGH 134 )
29X-RAY DIFFRACTION29CHAIN 'G' AND (RESID 135 THROUGH 323 )
30X-RAY DIFFRACTION30CHAIN 'H' AND (RESID 8 THROUGH 31 )
31X-RAY DIFFRACTION31CHAIN 'H' AND (RESID 32 THROUGH 58 )
32X-RAY DIFFRACTION32CHAIN 'H' AND (RESID 59 THROUGH 118 )
33X-RAY DIFFRACTION33CHAIN 'H' AND (RESID 119 THROUGH 218 )
34X-RAY DIFFRACTION34CHAIN 'H' AND (RESID 219 THROUGH 273 )
35X-RAY DIFFRACTION35CHAIN 'H' AND (RESID 274 THROUGH 333 )
36X-RAY DIFFRACTION36CHAIN 'H' AND (RESID 334 THROUGH 373 )
37X-RAY DIFFRACTION37CHAIN 'I' AND (RESID 2 THROUGH 48 )
38X-RAY DIFFRACTION38CHAIN 'I' AND (RESID 49 THROUGH 68 )
39X-RAY DIFFRACTION39CHAIN 'I' AND (RESID 69 THROUGH 148 )
40X-RAY DIFFRACTION40CHAIN 'I' AND (RESID 149 THROUGH 175 )
41X-RAY DIFFRACTION41CHAIN 'I' AND (RESID 176 THROUGH 230 )

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