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- PDB-8et0: Crystal Complex of murine Cyclooxygenase-2 with alpaca nanobody F9 -

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Basic information

Entry
Database: PDB / ID: 8et0
TitleCrystal Complex of murine Cyclooxygenase-2 with alpaca nanobody F9
Components
  • Prostaglandin G/H synthase 2
  • anti-cox-2 alpaca nanobody F9
KeywordsMEMBRANE PROTEIN / cyclooxygenase / nanobody / protein-antibody complex
Function / homology
Function and homology information


Biosynthesis of DHA-derived SPMs / Biosynthesis of EPA-derived SPMs / Biosynthesis of DPAn-3 SPMs / Biosynthesis of electrophilic ω-3 PUFA oxo-derivatives / Synthesis of 15-eicosatetraenoic acid derivatives / cellular response to non-ionic osmotic stress / positive regulation of platelet-derived growth factor production / hair cycle / cellular response to homocysteine / Nicotinamide salvaging ...Biosynthesis of DHA-derived SPMs / Biosynthesis of EPA-derived SPMs / Biosynthesis of DPAn-3 SPMs / Biosynthesis of electrophilic ω-3 PUFA oxo-derivatives / Synthesis of 15-eicosatetraenoic acid derivatives / cellular response to non-ionic osmotic stress / positive regulation of platelet-derived growth factor production / hair cycle / cellular response to homocysteine / Nicotinamide salvaging / Synthesis of Prostaglandins (PG) and Thromboxanes (TX) / positive regulation of fibroblast growth factor production / prostaglandin-endoperoxide synthase / prostaglandin-endoperoxide synthase activity / negative regulation of synaptic transmission, dopaminergic / cellular response to lead ion / response to nematode / positive regulation of transforming growth factor beta production / negative regulation of intrinsic apoptotic signaling pathway in response to osmotic stress / positive regulation of prostaglandin biosynthetic process / regulation of neuroinflammatory response / positive regulation of synaptic plasticity / positive regulation of smooth muscle contraction / oxidoreductase activity, acting on single donors with incorporation of molecular oxygen, incorporation of two atoms of oxygen / cyclooxygenase pathway / response to fatty acid / response to fructose / positive regulation of fever generation / prostaglandin secretion / response to vitamin D / cellular response to fluid shear stress / nuclear outer membrane / response to angiotensin / response to manganese ion / prostaglandin biosynthetic process / nuclear inner membrane / negative regulation of smooth muscle contraction / cellular response to ATP / positive regulation of cell migration involved in sprouting angiogenesis / maintenance of blood-brain barrier / bone mineralization / negative regulation of calcium ion transport / decidualization / negative regulation of cell cycle / positive regulation of vascular endothelial growth factor production / response to tumor necrosis factor / brown fat cell differentiation / response to glucocorticoid / keratinocyte differentiation / positive regulation of vasoconstriction / positive regulation of brown fat cell differentiation / embryo implantation / positive regulation of synaptic transmission, glutamatergic / learning / response to cytokine / caveola / positive regulation of smooth muscle cell proliferation / negative regulation of cysteine-type endopeptidase activity involved in apoptotic process / memory / regulation of blood pressure / positive regulation of protein import into nucleus / peroxidase activity / cellular response to mechanical stimulus / positive regulation of nitric oxide biosynthetic process / positive regulation of peptidyl-serine phosphorylation / response to estradiol / cellular response to heat / regulation of cell population proliferation / cellular response to hypoxia / angiogenesis / response to oxidative stress / response to lipopolysaccharide / response to xenobiotic stimulus / neuron projection / positive regulation of apoptotic process / negative regulation of cell population proliferation / heme binding / positive regulation of cell population proliferation / endoplasmic reticulum membrane / negative regulation of apoptotic process / enzyme binding / protein homodimerization activity / protein-containing complex / metal ion binding / cytoplasm
Similarity search - Function
Haem peroxidase, animal-type / Haem peroxidase domain superfamily, animal type / Animal haem peroxidase / Animal heme peroxidase superfamily profile. / EGF-like domain / Haem peroxidase superfamily / EGF-like domain profile. / EGF-like domain
Similarity search - Domain/homology
IBUPROFEN / Prostaglandin G/H synthase 2
Similarity search - Component
Biological speciesMus musculus (house mouse)
Vicugna pacos (alpaca)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.15 Å
AuthorsXu, S. / Banerjee, S. / Uddin, M.J. / Goodman, M.C. / Marnett, L.J.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM15431 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)CA089450 United States
CitationJournal: To Be Published
Title: Crystal complex of murine cycloxygenase-2 with alpaca nanobody F9
Authors: Xu, S. / Uddin, M.J.
History
DepositionOct 15, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 18, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Prostaglandin G/H synthase 2
B: Prostaglandin G/H synthase 2
C: anti-cox-2 alpaca nanobody F9
D: anti-cox-2 alpaca nanobody F9
hetero molecules


Theoretical massNumber of molelcules
Total (without water)167,30112
Polymers165,1554
Non-polymers2,1468
Water9,206511
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area13240 Å2
ΔGint6 kcal/mol
Surface area50380 Å2
MethodPISA
Unit cell
Length a, b, c (Å)141.526, 141.526, 90.429
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number76
Space group name H-MP41

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Components

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Protein / Antibody , 2 types, 4 molecules ABCD

#1: Protein Prostaglandin G/H synthase 2 / Cyclooxygenase-2 / COX-2 / Glucocorticoid-regulated inflammatory cyclooxygenase / Gripghs / ...Cyclooxygenase-2 / COX-2 / Glucocorticoid-regulated inflammatory cyclooxygenase / Gripghs / Macrophage activation-associated marker protein P71/73 / PES-2 / PHS II / Prostaglandin H2 synthase 2 / PGHS-2 / Prostaglandin-endoperoxide synthase 2 / TIS10 protein


Mass: 67332.711 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: apalca nanobody / Source: (gene. exp.) Mus musculus (house mouse) / Gene: Ptgs2, Cox-2, Cox2, Pghs-b, Tis10 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: Q05769, prostaglandin-endoperoxide synthase
#2: Antibody anti-cox-2 alpaca nanobody F9


Mass: 15244.560 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vicugna pacos (alpaca) / Production host: Escherichia coli (E. coli)

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Sugars , 2 types, 6 molecules

#3: Polysaccharide 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#4: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 2 types, 513 molecules

#5: Chemical ChemComp-IBP / IBUPROFEN / 2-(4-ISOBUTYLPHENYL)PROPIONIC ACID


Mass: 206.281 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C13H18O2 / Comment: antiinflammatory, medication*YM
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 511 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN

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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: 2.74 Å3/Da / Density % sol: 55.14 %
Crystal growTemperature: 291 K / Method: vapor diffusion, hanging drop / pH: 6.5
Details: 0.1 M Tris-Cl pH 8.5, 40-75% (+/-)-2-Methyl-2,4-pentanediol

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-C / Wavelength: 0.97918 Å
DetectorType: PSI PILATUS 6M / Detector: PIXEL / Date: Oct 19, 2017
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97918 Å / Relative weight: 1
ReflectionResolution: 2.147→141.53 Å / Num. obs: 190724 / % possible obs: 99.7 % / Redundancy: 9 % / CC1/2: 0.998 / Rmerge(I) obs: 0.106 / Rpim(I) all: 0.037 / Rrim(I) all: 0.112 / Net I/σ(I): 13.5 / Num. measured all: 877793 / Scaling rejects: 39
Reflection shellResolution: 2.15→2.18 Å / Redundancy: 8.7 % / Rmerge(I) obs: 2.098 / Num. measured all: 39944 / Num. unique obs: 4570 / CC1/2: 0.491 / Rpim(I) all: 0.734 / Rrim(I) all: 2.225 / Net I/σ(I) obs: 0.9 / % possible all: 94.4

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Processing

Software
NameVersionClassification
PHENIX1.12_2829refinement
Aimless0.5.23data scaling
PDB_EXTRACT3.27data extraction
XDSdata reduction
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 3NT1

3nt1


Resolution: 2.15→76.2 Å / SU ML: 0.3 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 23.4 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.214 5762 3.02 %
Rwork0.181 184927 -
obs0.182 190689 99.6 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 176.61 Å2 / Biso mean: 60.35 Å2 / Biso min: 31.97 Å2
Refinement stepCycle: final / Resolution: 2.15→76.2 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms10581 0 142 511 11234
Biso mean--76.13 55.82 -
Num. residues----1314
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection Rwork% reflection obs (%)
2.15-2.17170.34011790.3158565892
2.1717-2.19720.30111940.3136161100
2.1972-2.2240.31951900.2986134100
2.224-2.25220.34961920.30186192100
2.2522-2.28180.34171950.31616189100
2.2818-2.31310.31041920.29446174100
2.3131-2.34610.3341900.26796137100
2.3461-2.38120.29882010.25356231100
2.3812-2.41840.26861880.2476181100
2.4184-2.4580.27692000.24136188100
2.458-2.50040.27221960.23146228100
2.5004-2.54590.29271960.21786160100
2.5459-2.59480.26821940.22086218100
2.5948-2.64780.24861920.21886140100
2.6478-2.70540.22451970.20916223100
2.7054-2.76830.23721940.19776131100
2.7683-2.83760.22931850.20786226100
2.8376-2.91430.20721920.19756159100
2.9143-30.23771940.19236214100
3-3.09690.23671910.19786196100
3.0969-3.20760.24331940.18396101100
3.2076-3.3360.20781980.1816283100
3.336-3.48780.22431960.16996147100
3.4878-3.67170.1671920.15296123100
3.6717-3.90170.16561860.13756234100
3.9017-4.2030.17621840.14716178100
4.203-4.62590.18691980.13976213100
4.6259-5.29510.17061870.15256165100
5.2951-6.67070.24111830.19026172100
6.6707-76.20.17961920.1589617199
Refinement TLS params.

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

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
11.42310.1302-0.79890.7078-0.01813.5587-0.0494-0.30170.01570.2282-0.1473-0.1345-0.06940.34840.13250.4113-0.0493-0.0860.4477-0.00590.5138190.87-31.88722.997
27.8925-1.5734-5.00945.83861.14314.9908-0.16840.1227-0.44260.0494-0.0203-0.46080.35270.27760.13280.30770.0356-0.06460.4462-0.02760.4659190.107-48.5910.653
31.73891.149-0.85232.5128-1.2522.24310.0865-0.04870.4040.286-0.05080.156-0.2364-0.0186-0.02660.40830.0131-0.04640.3759-0.05820.4671173.71-23.717.876
40.82490.26110.0781.3776-0.41060.9624-0.0050.20080.1759-0.28150.02650.2640.1338-0.14570.00750.43830.0164-0.10650.437-0.02330.4228160.992-36.681-2.691
51.8644-1.403-0.27468.1683-1.44121.3903-0.03880.38380.0296-0.45140.09070.70640.0788-0.1896-0.04570.5063-0.0556-0.18570.526-0.02060.404155.424-39.867-8.032
61.32410.18260.20741.5031-0.09220.7124-0.00920.25150.1956-0.2926-0.03550.00660.01190.06550.04750.38460.0099-0.03420.41030.01550.3386175.591-32.022-0.688
71.87020.4704-0.18365.0233-3.38298.2356-0.13930.3345-0.2823-0.6999-0.0694-0.09880.41220.0910.16620.41430.0649-0.05810.3845-0.09830.3827170.104-51.838-6.095
80.93020.13060.04411.65840.28042.1733-0.1185-0.014-0.2104-0.2255-0.0603-0.01750.21310.07880.14820.46940.0131-0.03150.3470.02410.5896171.126-72.0420.186
97.3246-0.88516.45442.3355-1.62616.91910.25980.2352-0.2059-0.0791-0.1725-0.01410.32590.3264-0.14180.37760.0342-0.01840.3309-0.03290.3616170.634-59.2123.53
101.11460.4399-0.47272.3314-1.58133.34090.03040.0482-0.1314-0.07880.06810.40350.2509-0.5048-0.1110.3461-0.0149-0.08090.4393-0.02970.5785149.45-65.6119.625
111.69380.2823-0.18162.3817-0.30971.05950.0877-0.35550.19420.3437-0.07480.2433-0.2108-0.153-0.01270.3815-0.00290.03510.4445-0.10270.3559154.495-44.84937.965
125.50631.9918-0.3024.2852-0.54232.11110.0422-0.50290.48830.4659-0.01730.6562-0.2738-0.1229-0.04020.55470.03170.09920.5051-0.13320.4197153.736-38.2342.99
131.2270.3478-0.02071.6057-0.05180.78570.0711-0.3285-0.12350.2735-0.07450.24560.0519-0.09870.00520.4098-0.0233-0.00410.4260.01310.3724158.743-59.71536.942
146.31722.4525-4.37933.2779-2.38147.3620.2257-0.7305-0.06560.5042-0.2442-0.1134-0.27230.1883-0.02560.4174-0.0031-0.08950.4314-0.02670.3299171.929-44.0542.716
154.88620.2799-6.12290.0171-0.34947.67730.26420.9285-1.1223-0.0287-1.56254.0950.0912-1.52391.42410.48490.008-0.0860.701-0.05461.1748145.796-16.8274.306
169.30653.91945.091.78051.71144.21140.1145-0.13350.0951-0.7528-0.7771.0852-1.6728-0.69040.55031.38690.28280.44650.8103-0.25491.5464143.7694.4812.914
173.41432.68121.82232.7895-0.55856.86510.6959-0.54980.00250.3533-0.30260.1514-0.2373-0.922-0.39080.44780.00730.02720.621-0.05230.9165147.364-14.46310.172
185.6244-5.67370.0387.316-1.65724.9177-0.09710.10380.5122-0.46850.1568-0.0341-0.1379-0.0505-0.10130.4381-0.0085-0.080.4316-0.07150.6739155.319-13.7464.923
199.3072-4.79744.40238.2262-5.63664.0490.49460.94710.3811-1.04270.07520.4847-0.074-0.15-0.66110.92020.0904-0.09480.56140.02090.7489155.899-3.2251.418
204.5508-4.82695.32355.1872-5.7326.331-0.41390.36990.47390.1762-0.39960.5218-1.81970.90920.72250.8027-0.1528-0.18480.4912-0.06960.6357162.884-10.71211.811
219.77131.7695-5.78917.6346-1.76563.51170.1157-0.13510.86030.7770.7204-0.859-0.53420.1832-0.75191.0912-0.2429-0.24510.50950.06730.7724160.2682.02210.703
226.13714.01140.55618.30371.87137.34310.2541-0.96230.2681.8034-0.43370.8643-0.6518-0.64870.14850.8771-0.04020.08080.5546-0.07290.7054151.858-9.84714.983
237.2686-2.7772-3.18986.92681.86712.5371.17480.24250.61242.18660.10871.4049-1.9204-0.21-1.10421.51320.16580.22340.53160.09331.2467149.6455.7956.488
247.7596-6.9604-5.12068.96385.97616.38910.27140.02390.0619-0.5833-0.07860.7935-0.606-0.3672-0.22330.49860.0658-0.08710.4571-0.00440.7132152.512-12.5251.022
254.6351-0.47454.79392.2196-1.08235.1629-0.9962-0.83855.0004-0.046-0.7650.8433-1.4204-2.2261.77420.98820.2522-0.0781.13720.01531.2785130.065-43.31628.975
260.0843-0.5558-0.60843.65464.03094.989-0.5322-0.33051.4126-0.28030.27970.9457-0.07920.28090.30410.7122-0.1002-0.2492.2964-0.46351.443111.938-54.17419.227
276.28871.75980.27296.5301-1.08560.2780.20750.6847-0.5369-0.0359-0.05311.5004-0.3879-2.3071-0.07880.50210.0734-0.03681.3512-0.12531.0258131.649-50.927.019
284.0228-1.5183-3.35472.852-1.17115.3942-0.28690.0029-0.74440.49480.59550.79270.2954-0.8229-0.30330.722-0.28790.28281.6901-0.5031.5291123.998-59.24332.144
292.45240.92070.48790.35810.17090.08980.14590.7001-1.36620.0299-0.63031.19811.2289-1.81320.31090.8609-0.46450.0551.8252-0.65191.6805128.24-63.88722.967
301.9908-0.76410.79137.58680.08426.4688-0.96532.34650.5469-1.18660.01781.72880.3949-2.71440.90950.8357-0.0031-0.10651.9682-0.17480.9647129.661-51.92818.35
310.5158-0.17990.81751.28160.36722.1627-0.09431.1696-1.4305-0.37920.66920.09690.308-0.5205-0.26810.784-0.30590.19712.6998-0.87722.2196113.73-59.7825.314
323.7045-0.08074.27733.8082-2.50846.4940.22231.0670.28750.0057-0.73831.44310.1317-1.07470.48240.47580.01290.0121.2826-0.19111.1237131.624-52.59931.46
337.15211.73382.27070.95830.37190.86840.04540.340.13170.1636-0.47131.4147-0.1063-1.6597-0.02190.4610.09120.05351.5689-0.27351.281128.871-50.5733.244
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1( CHAIN A AND RESID 33:93 )A33 - 93
2X-RAY DIFFRACTION2( CHAIN A AND RESID 94:123 )A94 - 123
3X-RAY DIFFRACTION3( CHAIN A AND RESID 124:181 )A124 - 181
4X-RAY DIFFRACTION4( CHAIN A AND RESID 182:269 )A182 - 269
5X-RAY DIFFRACTION5( CHAIN A AND RESID 270:319 )A270 - 319
6X-RAY DIFFRACTION6( CHAIN A AND RESID 320:535 )A320 - 535
7X-RAY DIFFRACTION7( CHAIN A AND RESID 536:583 )A536 - 583
8X-RAY DIFFRACTION8( CHAIN B AND RESID 33:105 )B33 - 105
9X-RAY DIFFRACTION9( CHAIN B AND RESID 106:138 )B106 - 138
10X-RAY DIFFRACTION10( CHAIN B AND RESID 139:181 )B139 - 181
11X-RAY DIFFRACTION11( CHAIN B AND RESID 182:269 )B182 - 269
12X-RAY DIFFRACTION12( CHAIN B AND RESID 270:319 )B270 - 319
13X-RAY DIFFRACTION13( CHAIN B AND RESID 320:535 )B320 - 535
14X-RAY DIFFRACTION14( CHAIN B AND RESID 536:583 )B536 - 583
15X-RAY DIFFRACTION15( CHAIN C AND RESID 3:9 )C3 - 9
16X-RAY DIFFRACTION16( CHAIN C AND RESID 10:20 )C10 - 20
17X-RAY DIFFRACTION17( CHAIN C AND RESID 21:29 )C21 - 29
18X-RAY DIFFRACTION18( CHAIN C AND RESID 30:41 )C30 - 41
19X-RAY DIFFRACTION19( CHAIN C AND RESID 42:54 )C42 - 54
20X-RAY DIFFRACTION20( CHAIN C AND RESID 55:61 )C55 - 61
21X-RAY DIFFRACTION21( CHAIN C AND RESID 62:68 )C62 - 68
22X-RAY DIFFRACTION22( CHAIN C AND RESID 69:86 )C69 - 86
23X-RAY DIFFRACTION23( CHAIN C AND RESID 87:94 )C87 - 94
24X-RAY DIFFRACTION24( CHAIN C AND RESID 95:118 )C95 - 118
25X-RAY DIFFRACTION25( CHAIN D AND RESID 3:9 )D3 - 9
26X-RAY DIFFRACTION26( CHAIN D AND RESID 10:20 )D10 - 20
27X-RAY DIFFRACTION27( CHAIN D AND RESID 21:41 )D21 - 41
28X-RAY DIFFRACTION28( CHAIN D AND RESID 43:54 )D43 - 54
29X-RAY DIFFRACTION29( CHAIN D AND RESID 55:69 )D55 - 69
30X-RAY DIFFRACTION30( CHAIN D AND RESID 70:82 )D70 - 82
31X-RAY DIFFRACTION31( CHAIN D AND RESID 88:94 )D88 - 94
32X-RAY DIFFRACTION32( CHAIN D AND RESID 95:103 )D95 - 103
33X-RAY DIFFRACTION33( CHAIN D AND RESID 104:118 )D104 - 118

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