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- PDB-3mb2: Kinetic and Structural Characterization of a Heterohexamer 4-Oxal... -

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Basic information

Entry
Database: PDB / ID: 3mb2
TitleKinetic and Structural Characterization of a Heterohexamer 4-Oxalocrotonate Tautomerase from Chloroflexus aurantiacus J-10-fl: Implications for Functional and Structural Diversity in the Tautomerase Superfamily
Components
  • 4-oxalocrotonate tautomerase family enzyme - alpha subunit
  • 4-oxalocrotonate tautomerase family enzyme - beta subunit
KeywordsISOMERASE / TRANS-3-CHLOROACRYLIC ACID DEHALOGENASE / CAAD / DEHALOGENASE / HYDROLASE / 4-Oxalocrotonate tautomerase / 4-OT / heterohexamer
Function / homology
Function and homology information


: / isomerase activity
Similarity search - Function
4-oxalocrotonate tautomerase / Tautomerase enzyme / Macrophage Migration Inhibitory Factor / Macrophage Migration Inhibitory Factor / Tautomerase/MIF superfamily / 2-Layer Sandwich / Alpha Beta
Similarity search - Domain/homology
4-oxalocrotonate tautomerase family enzyme / Uncharacterized protein
Similarity search - Component
Biological speciesChloroflexus aurantiacus (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.41 Å
AuthorsBurks, E.A. / Fleming, C.D. / Mesecar, A.D. / Whitman, C.P. / Pegan, S.D.
CitationJournal: Biochemistry / Year: 2010
Title: Kinetic and structural characterization of a heterohexamer 4-oxalocrotonate tautomerase from Chloroflexus aurantiacus J-10-fl: implications for functional and structural diversity in the tautomerase superfamily
Authors: Burks, E.A. / Fleming, C.D. / Mesecar, A.D. / Whitman, C.P. / Pegan, S.D.
History
DepositionMar 24, 2010Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 2, 2010Provider: repository / Type: Initial release
Revision 1.1Jul 13, 2011Group: Advisory / Source and taxonomy / Version format compliance
Revision 1.2Sep 6, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
B: 4-oxalocrotonate tautomerase family enzyme - beta subunit
C: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
D: 4-oxalocrotonate tautomerase family enzyme - beta subunit
E: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
F: 4-oxalocrotonate tautomerase family enzyme - beta subunit
G: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
H: 4-oxalocrotonate tautomerase family enzyme - beta subunit
I: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
J: 4-oxalocrotonate tautomerase family enzyme - beta subunit
K: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
L: 4-oxalocrotonate tautomerase family enzyme - beta subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)95,89829
Polymers94,26512
Non-polymers1,63317
Water4,486249
1
A: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
B: 4-oxalocrotonate tautomerase family enzyme - beta subunit
C: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
D: 4-oxalocrotonate tautomerase family enzyme - beta subunit
E: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
F: 4-oxalocrotonate tautomerase family enzyme - beta subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)47,99715
Polymers47,1326
Non-polymers8659
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area16170 Å2
ΔGint-257 kcal/mol
Surface area14580 Å2
MethodPISA
2
G: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
H: 4-oxalocrotonate tautomerase family enzyme - beta subunit
I: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
J: 4-oxalocrotonate tautomerase family enzyme - beta subunit
K: 4-oxalocrotonate tautomerase family enzyme - alpha subunit
L: 4-oxalocrotonate tautomerase family enzyme - beta subunit
hetero molecules


Theoretical massNumber of molelcules
Total (without water)47,90114
Polymers47,1326
Non-polymers7698
Water1086
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area15560 Å2
ΔGint-201 kcal/mol
Surface area14580 Å2
MethodPISA
Unit cell
Length a, b, c (Å)106.062, 106.062, 109.961
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number144
Space group name H-MP31
DetailsBiological assembly is one of the two heterohexamers located in the asymmetric unit.

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Components

#1: Protein
4-oxalocrotonate tautomerase family enzyme - alpha subunit


Mass: 7738.895 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chloroflexus aurantiacus (bacteria) / Strain: J-10-fl / Gene: 4-oxalocrotonate tautomerase, Caur_1354 / Plasmid: pET24a / Production host: Escherichia coli (E. coli) / Strain (production host): BL21-Gold(DE3), / References: UniProt: A9W9U6, EC: 5.3.2.2
#2: Protein
4-oxalocrotonate tautomerase family enzyme - beta subunit


Mass: 7971.898 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Chloroflexus aurantiacus (bacteria) / Strain: J-10-fl / Gene: 4-oxalocrotonate tautomerase, Caur_1358 / Plasmid: pET24a / Production host: Escherichia coli (E. coli) / Strain (production host): BL21-Gold(DE3), / References: UniProt: A9W9V0, EC: 5.3.2.2
#3: Chemical
ChemComp-SO4 / SULFATE ION


Mass: 96.063 Da / Num. of mol.: 17 / Source method: obtained synthetically / Formula: SO4
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 249 / 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: 3.79 Å3/Da / Density % sol: 67.53 %
Crystal growTemperature: 293 K / Method: vapor diffusion, hanging drop / pH: 7.4
Details: 0.25 M (NH4)2SO4 and 4% PEG 4000, pH 7.4, VAPOR DIFFUSION, HANGING DROP, temperature 293K

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 22-BM / Wavelength: 1 Å
DetectorType: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Apr 16, 2008
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 2.41→91.85 Å / Num. all: 49937 / Num. obs: 49737 / % possible obs: 99.6 % / Redundancy: 6 % / Biso Wilson estimate: 32.143 Å2 / Rmerge(I) obs: 0.072 / Net I/σ(I): 27.6
Reflection shellResolution: 2.414→2.477 Å / Redundancy: 5.2 % / Rmerge(I) obs: 0.377 / Mean I/σ(I) obs: 4.1 / Num. unique all: 3293 / % possible all: 88.69

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Processing

Software
NameVersionClassification
HKL-2000data collection
PHASERphasing
REFMAC5.5.0102refinement
HKL-2000data reduction
HKL-2000data scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB ENTRY 1S0Y Alpha Subunit PDB ENTRY 1MWW Beta Subunit
Resolution: 2.41→91.85 Å / Cor.coef. Fo:Fc: 0.952 / Cor.coef. Fo:Fc free: 0.928 / SU B: 14.171 / SU ML: 0.154 / TLS residual ADP flag: LIKELY RESIDUAL / Isotropic thermal model: Isotropic / Cross valid method: THROUGHOUT / ESU R: 0.234 / ESU R Free: 0.211 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.25018 2670 5.1 %RANDOM
Rwork0.20324 ---
obs0.20556 49737 98.58 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.4 Å / Solvent model: MASK
Displacement parametersBiso mean: 32.143 Å2
Baniso -1Baniso -2Baniso -3
1-0.44 Å20.22 Å20 Å2
2--0.44 Å2-0 Å2
3----0.67 Å2
Refinement stepCycle: LAST / Resolution: 2.41→91.85 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5422 0 85 249 5756
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0260.0225608
X-RAY DIFFRACTIONr_angle_refined_deg2.2262.0027629
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.5295703
X-RAY DIFFRACTIONr_dihedral_angle_2_deg39.88123220
X-RAY DIFFRACTIONr_dihedral_angle_3_deg22.16715947
X-RAY DIFFRACTIONr_dihedral_angle_4_deg20.1491556
X-RAY DIFFRACTIONr_chiral_restr0.1640.2904
X-RAY DIFFRACTIONr_gen_planes_refined0.0110.0214088
X-RAY DIFFRACTIONr_mcbond_it0.9281.53569
X-RAY DIFFRACTIONr_mcangle_it1.72125763
X-RAY DIFFRACTIONr_scbond_it3.32632039
X-RAY DIFFRACTIONr_scangle_it5.14.51866
LS refinement shellResolution: 2.414→2.477 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.407 182 -
Rwork0.301 3293 -
obs--88.69 %
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)
13.24761.4579-1.18035.4276-2.84976.23040.19880.13650.1366-0.0206-0.328-0.0099-0.29910.1690.12920.06720.02720.00350.0475-0.00940.06128.868515.9133-5.5961
25.41612.7818-1.3145.10910.19662.06040.2277-0.35050.12790.2306-0.42060.3203-0.0447-0.08680.19290.09790.02960.01030.0734-0.03510.03676.08711.3389-1.1528
35.9662-3.94960.33414.7972-1.65565.9745-0.29080.31290.7822-0.92450.0248-0.9396-0.16370.52540.2660.322-0.00760.01230.31240.04240.226829.03817.4508-6.2004
44.2750.11532.03787.2809-0.17574.45810.10980.0657-0.54890.1445-0.0383-0.46080.39280.3885-0.07150.04740.0073-0.01640.1360.05120.121429.08333.674710.7911
55.10083.16542.25775.58463.55244.74260.3221-0.13810.04480.5206-0.2762-0.26950.2080.1949-0.04590.0724-0.0299-0.02680.08190.05190.072226.68510.834211.894
615.613620.360514.500919.036216.988911.96750.77350.0018-1.46280.67640.0806-1.50570.74760.0456-0.85410.74980.34980.43330.38890.03170.682417.3988-7.9941-5.7068
73.7813-2.20750.7385.55240.35534.62030.1478-0.04070.2399-0.086-0.14950.1573-0.29090.00630.00170.0795-0.06170.01490.0672-0.01060.04158.404224.330715.8373
82.81230.65181.55171.8045-0.51965.98130.08140.0850.00720.0373-0.3231-0.1799-0.14380.61640.24170.0216-0.02150.02270.10540.03230.090510.333918.662716.4413
919.3007-11.864310.588226.0661-5.30845.6011-0.9569-0.91280.3660.66360.74211.9202-0.6039-0.74060.21480.41290.29750.00860.81620.17630.6324-4.682523.0382-2.6377
102.1684-2.30391.18823.83771.27496.1879-0.1252-0.0834-0.18810.5951-0.02540.0990.4774-0.17320.15070.2014-0.0183-0.09920.06350.01130.15087.36920.28193.7357
111.22440.1561-1.76762.7727-0.80594.92890.0905-0.0833-0.0537-0.2829-0.2395-0.08290.21150.42670.14890.07260.05180.00130.05530.00380.042911.6173.9689-3.5006
1210.8011-1.4406-10.2939-0.11371.027320.88050.0719-0.3373-0.8423-0.00030.120.18310.1714-0.9735-0.19190.1277-0.06030.04480.22380.06810.362-1.15384.589119.386
132.8701-1.2438-1.08986.1141-0.81692.56460.0195-0.15740.1821-0.1072-0.2213-0.0441-0.32670.30580.20180.08280.0286-0.04060.20350.04290.058926.331920.07396.0912
141.37840.0614-0.47172.93450.20182.56130.1490.1477-0.067-0.3957-0.209-0.2941-0.02620.26180.05990.11650.0092-0.020.12380.0710.100627.786416.13336.167
1517.22677.53060.85843.2602-4.33324.22420.4326-0.29370.82580.5986-0.37590.2767-1.77021.0295-0.05670.5084-0.2152-0.04780.2250.03740.598519.035231.450122.0017
169.115-0.3055-2.11083.10961.50582.03110.1068-0.6917-0.697-0.0661-0.1871-0.38690.52340.26870.08030.35790.10860.06650.13830.08510.117311.82217.155222.1875
17-0.0279-0.4931-0.609515.7344-5.85466.567-0.0468-0.036-0.01140.47080.27590.532-0.0183-0.0007-0.22910.0597-0.03590.03370.0591-0.00790.03522.25913.601723.3504
188.5002-4.3751-0.20634.1568-0.59431.42540.103-0.0494-0.32290.0962-0.0950.19550.08230.1118-0.0080.1015-0.0457-0.02690.03620.01530.01959.359811.548916.8261
193.4569-3.12411.33246.5165-3.34415.09910.081-0.0402-0.0850.1595-0.214-0.11140.23210.2290.13290.0759-0.03530.01450.0553-0.02950.07919.624345.03557.8404
206.4198-2.65160.91385.1462-0.55551.66830.16940.3245-0.1131-0.0475-0.34680.38910.0492-0.030.17740.1034-0.04930.00180.0609-0.03450.0565.778249.65623.991
210.3835-0.0068-1.71766.8815-6.32212.0904-0.2563-0.228-0.14861.5234-0.2381-0.3938-0.14270.95560.49440.5776-0.017-0.11010.34130.10410.333729.082843.74768.6824
224.0517-1.7901-2.10594.7709-1.35484.17620.212-0.04270.6047-0.2489-0.018-0.2052-0.48830.4455-0.1940.1465-0.0851-0.00080.18680.01430.128928.303858.3315-7.9362
237.2715-3.4371-0.76027.27791.72194.69390.13640.1785-0.087-0.644-0.0536-0.65270.07940.2541-0.08280.10110.01070.08240.09250.05980.140231.080845.8018-12.9786
245.2655-0.7624-4.6597.36052.270410.2538-0.0583-0.4580.581-0.008-0.0282-0.3269-1.1140.31060.08650.2526-0.0103-0.10340.0932-0.06690.174817.213965.14422.637
253.77582.2456-1.37715.8126-0.67555.74660.20720.1057-0.32130.1379-0.23920.17450.2622-0.00050.03190.07340.0522-0.00820.0673-0.01750.06388.323636.9068-13.5783
262.2958-0.3565-0.67481.2578-0.275.45740.1361-0.0303-0.0027-0.0277-0.3157-0.1442-0.07880.42990.17960.01730.0039-0.02570.10610.02610.095710.704542.4652-14.58
2713.8011-2.30964.308131.4453-2.0554.70520.117-0.5116-0.95980.4517-0.11361.02310.9588-0.5836-0.00340.2827-0.11130.08680.16580.03140.2164-2.499741.21045.2655
283.01323.1646-0.69554.78090.97154.951-0.05870.20250.323-0.5599-0.35780.2395-0.4556-0.02340.41650.19340.13780.04250.33230.1410.13887.673660.3375-1.3409
291.18830.40210.95082.0555-0.47134.17580.08310.03330.02010.1842-0.2745-0.0866-0.1960.29650.19150.0434-0.03910.0090.0574-0.00030.060910.974557.43814.8959
302.3286-3.714-1.89655.12273.5811.38180.28170.1186-0.2421-0.3742-0.26470.4695-0.6263-1.4737-0.0170.14450.06670.04220.28230.01410.4847-3.388859.0883-17.9752
314.3763-0.8820.51549.28840.73542.1557-0.074-0.0723-0.2810.1595-0.0495-0.32760.24970.29960.12350.0905-0.02710.02330.15970.0530.058728.190940.7123-3.3402
322.59251.33120.57122.88050.6953.74080.3189-0.28430.12440.4153-0.3251-0.18930.03110.16260.00620.1114-0.01210.02540.11750.06540.13726.827246.1043-3.8527
3310.3786-11.3224-0.191511.47086.483730.0210.30750.5398-0.3729-0.0692-0.16670.15691.45871.2048-0.14070.4937-0.01380.10720.3606-0.20740.642719.776828.9196-19.7598
346.16640.24152.9043.10980.9641.8437-0.20990.59930.5784-0.0996-0.0683-0.3412-0.41250.21680.27820.30970.03-0.11090.06590.0490.128511.694853.8543-19.9218
353.41072.3731-0.29636.0351-1.2014.41110.0578-0.02030.1076-0.0570.07830.3771-0.0134-0.1059-0.1360.03370.0243-0.01470.0215-0.00190.03552.08846.8135-16.668
3617.2529-4.57674.643719.121-3.25823.65860.12271.0275-0.229-0.7835-0.3703-0.5181-0.54640.3580.24750.25770.0143-0.0110.12210.05330.111424.730357.0145-16.7565
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A1 - 22
2X-RAY DIFFRACTION2A23 - 50
3X-RAY DIFFRACTION3A51 - 61
4X-RAY DIFFRACTION4B1 - 24
5X-RAY DIFFRACTION5B25 - 53
6X-RAY DIFFRACTION6B54 - 59
7X-RAY DIFFRACTION7C1 - 22
8X-RAY DIFFRACTION8C23 - 55
9X-RAY DIFFRACTION9C56 - 62
10X-RAY DIFFRACTION10D1 - 16
11X-RAY DIFFRACTION11D17 - 51
12X-RAY DIFFRACTION12D52 - 58
13X-RAY DIFFRACTION13E1 - 15
14X-RAY DIFFRACTION14E16 - 55
15X-RAY DIFFRACTION15E56 - 61
16X-RAY DIFFRACTION16F1 - 16
17X-RAY DIFFRACTION17F17 - 30
18X-RAY DIFFRACTION18F31 - 54
19X-RAY DIFFRACTION19G1 - 20
20X-RAY DIFFRACTION20G21 - 50
21X-RAY DIFFRACTION21G51 - 61
22X-RAY DIFFRACTION22H1 - 21
23X-RAY DIFFRACTION23H22 - 45
24X-RAY DIFFRACTION24H46 - 58
25X-RAY DIFFRACTION25I1 - 22
26X-RAY DIFFRACTION26I23 - 53
27X-RAY DIFFRACTION27I54 - 60
28X-RAY DIFFRACTION28J1 - 15
29X-RAY DIFFRACTION29J16 - 53
30X-RAY DIFFRACTION30J54 - 59
31X-RAY DIFFRACTION31K1 - 20
32X-RAY DIFFRACTION32K21 - 55
33X-RAY DIFFRACTION33K56 - 62
34X-RAY DIFFRACTION34L1 - 16
35X-RAY DIFFRACTION35L17 - 46
36X-RAY DIFFRACTION36L47 - 54

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