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- PDB-8a8h: APS kinase from Methanothermococcus thermolithotrophicus refined ... -

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Basic information

Entry
Database: PDB / ID: 8a8h
TitleAPS kinase from Methanothermococcus thermolithotrophicus refined to 1.77 A
ComponentsAPS kinase from Methanothermococcus thermolithotrophicus
KeywordsTRANSFERASE / sulfate assimilation / methanogens / archaea / APS / ATP / ADP / PAPS / thermophile / methane / activation / marine
Function / homologyP-loop containing nucleotide triphosphate hydrolases / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta / PHOSPHATE ION
Function and homology information
Biological speciesMethanothermococcus thermolithotrophicus DSM 2095 (archaea)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.77 Å
AuthorsJespersen, M. / Wagner, T.
Funding support Germany, 2items
OrganizationGrant numberCountry
Max Planck Society Germany
German Research Foundation (DFG)WA 4053/1-1 Germany
CitationJournal: Nat Microbiol / Year: 2023
Title: Assimilatory sulfate reduction in the marine methanogen Methanothermococcus thermolithotrophicus.
Authors: Jespersen, M. / Wagner, T.
History
DepositionJun 23, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 26, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 15, 2023Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / citation
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
Revision 1.2Feb 7, 2024Group: Refinement description / Category: pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: APS kinase from Methanothermococcus thermolithotrophicus
B: APS kinase from Methanothermococcus thermolithotrophicus
C: APS kinase from Methanothermococcus thermolithotrophicus
D: APS kinase from Methanothermococcus thermolithotrophicus
hetero molecules


Theoretical massNumber of molelcules
Total (without water)88,33828
Polymers86,8234
Non-polymers1,51524
Water10,917606
1
A: APS kinase from Methanothermococcus thermolithotrophicus
B: APS kinase from Methanothermococcus thermolithotrophicus
hetero molecules


Theoretical massNumber of molelcules
Total (without water)44,16914
Polymers43,4122
Non-polymers75712
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4010 Å2
ΔGint-21 kcal/mol
Surface area14410 Å2
MethodPISA
2
C: APS kinase from Methanothermococcus thermolithotrophicus
D: APS kinase from Methanothermococcus thermolithotrophicus
hetero molecules


Theoretical massNumber of molelcules
Total (without water)44,16914
Polymers43,4122
Non-polymers75712
Water362
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area4130 Å2
ΔGint-22 kcal/mol
Surface area14390 Å2
MethodPISA
Unit cell
Length a, b, c (Å)51.539, 176.176, 51.610
Angle α, β, γ (deg.)90.000, 105.780, 90.000
Int Tables number4
Space group name H-MP1211

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Components

#1: Protein
APS kinase from Methanothermococcus thermolithotrophicus


Mass: 21705.867 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Details: The gene construct was expressed in the pET-28a(+) vector. The protein product contained a His-tag in its N-terminal part and a thrombin cleavage site in-between. The tag was cleaved off ...Details: The gene construct was expressed in the pET-28a(+) vector. The protein product contained a His-tag in its N-terminal part and a thrombin cleavage site in-between. The tag was cleaved off during the purification by the thrombin.
Source: (gene. exp.) Methanothermococcus thermolithotrophicus DSM 2095 (archaea)
Strain: DSM 2095 / Tissue: / / Cell: / / Cell line: / / Gene: cysC / Organ: / / Variant: / / Plasmid: pET-28a(+)
Details (production host): The synthetic gene was cloned in pET-28a(+) by using the restriction sites NdeI and BamHI. A stop codon (TGA) was incorporated before BamHI
Cell (production host): / / Cell line (production host): / / Organ (production host): / / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / Tissue (production host): / / Variant (production host): / / References: adenylyl-sulfate kinase
#2: Chemical
ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL


Mass: 62.068 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: C2H6O2
#3: Chemical
ChemComp-PO4 / PHOSPHATE ION


Mass: 94.971 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: PO4 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical
ChemComp-CL / CHLORIDE ION


Mass: 35.453 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Cl
#5: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 606 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestY

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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: 2.6 Å3/Da / Density % sol: 52.73 % / Description: Transparent, plate-shaped crystals
Crystal growTemperature: 291 K / Method: vapor diffusion, sitting drop / pH: 5.5
Details: MtAPSK (0.7 ul) with 2 mM MgCl2 at a protein concentration of 17.6 mg.ml-1 was mixed with 0.7 ul reservoir solution. Crystals were obtained in the following crystallization condition: 20 % ...Details: MtAPSK (0.7 ul) with 2 mM MgCl2 at a protein concentration of 17.6 mg.ml-1 was mixed with 0.7 ul reservoir solution. Crystals were obtained in the following crystallization condition: 20 % w/v polyethylene glycol 3350 and 100 mM tri-Sodium citrate pH 5.5. The cryoprotectant used for the crystals was 20 % w/v polyethylene glycol 3350, 100 mM tri-Sodium citrate pH 5.5 and 25% v/v ethylene glycol.
PH range: / / Temp details: 291 +/- 1 K

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X06DA / Wavelength: 1.00003 Å
DetectorType: DECTRIS PILATUS 2M-F / Detector: PIXEL / Date: Sep 29, 2019
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.00003 Å / Relative weight: 1
ReflectionResolution: 1.77→47.74 Å / Num. obs: 68557 / % possible obs: 83.1 % / Redundancy: 6.8 % / Biso Wilson estimate: 28.8 Å2 / CC1/2: 1 / Rmerge(I) obs: 0.044 / Rpim(I) all: 0.018 / Rrim(I) all: 0.048 / Net I/σ(I): 22.7
Reflection shellResolution: 1.77→1.81 Å / Redundancy: 6.2 % / Rmerge(I) obs: 0.928 / Mean I/σ(I) obs: 1.7 / Num. unique obs: 3427 / CC1/2: 0.685 / Rpim(I) all: 0.398 / Rrim(I) all: 1.001 / % possible all: 96.3

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Processing

Software
NameVersionClassification
PHENIX1.19.2_4158refinement
PDB_EXTRACT3.27data extraction
autoPROCdata reduction
autoPROCdata scaling
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5CB6

5cb6


Resolution: 1.77→44.04 Å / SU ML: 0.21 / Cross valid method: THROUGHOUT / σ(F): 1.35 / Phase error: 19.53 / Stereochemistry target values: ML
Details: The final refinement cycle was performed without non-crystallography symmetry and with translational-liberation screw. Hydrogens were omitted from the final refinement, but used in riding ...Details: The final refinement cycle was performed without non-crystallography symmetry and with translational-liberation screw. Hydrogens were omitted from the final refinement, but used in riding position during previous cycles.
RfactorNum. reflection% reflection
Rfree0.1809 3227 4.71 %
Rwork0.154 65324 -
obs0.1553 68551 79.96 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 107.21 Å2 / Biso mean: 37.1121 Å2 / Biso min: 16.76 Å2
Refinement stepCycle: final / Resolution: 1.77→44.04 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms4698 0 88 606 5392
Biso mean--49.79 42.68 -
Num. residues----586
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 23

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
1.77-1.80.30561060.282217232363
1.8-1.820.2941140.25672438255269
1.82-1.850.29751220.23432671279375
1.85-1.890.255890.23531826191552
1.89-1.90.2985900.25341555164584
1.93-1.960.271400.274759463420
1.96-20.24711530.21453384353795
2-2.040.25941880.226435103698100
2.04-2.090.2748450.234483888324
2.09-2.140.19762160.1635253741100
2.14-2.20.19621780.146635023680100
2.2-2.260.19691100.15462345245565
2.26-2.340.18331760.138935563732100
2.34-2.420.16591470.143435393686100
2.42-2.520.17541570.148735523709100
2.52-2.630.19711900.153935213711100
2.63-2.770.17561170.14472537265471
2.77-2.940.15241840.142735403724100
2.94-3.170.1671600.153535923752100
3.17-3.490.16291670.14283035320286
3.49-3.990.16791340.14232875300980
4-5.030.16581740.125435573731100
5.03-44.040.17291700.166936153785100
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)
19.53391.62082.65092.24921.10995.4555-0.0129-0.32140.21420.07410.1278-0.3991-0.30820.7162-0.21120.2986-0.0508-0.0120.3053-0.01570.214125.41322.7902-36.1112
23.25521.41482.09492.93372.00598.11220.1181-0.1733-0.16520.2282-0.0284-0.05880.6206-0.1782-0.17420.261-0.0329-0.01630.20410.00150.274819.3777-8.8889-36.7909
38.5987-1.78830.43914.19921.34725.05140.169-0.11410.16750.2002-0.15460.180.3924-0.47030.00720.2875-0.09060.00730.29480.02690.146814.7593-1.355-29.7643
43.6333-2.98564.02576.3043-4.05264.56540.00390.3311-0.529-0.20080.07881.69290.2855-1.5207-0.30450.31070.017-0.03680.8138-0.06310.4658-0.31255.4078-33.4815
58.4218-0.85352.71573.8269-0.4855.9454-0.02330.48480.3833-0.1601-0.0640.1987-0.3089-0.46510.00970.32590.03540.05010.34830.00030.163212.89739.8517-34.2485
61.7824-0.1461-0.26342.2257-0.76695.1912-0.0146-0.05310.0184-0.06780.073-0.0552-0.3024-0.2022-0.03710.2464-0.02520.02070.2479-0.00940.167318.74374.7405-42.2127
76.2549-1.6919-0.03073.6022-1.18424.4192-0.07940.5226-0.5464-0.42310.02570.23540.2316-0.47490.05380.3142-0.0552-0.00120.2973-0.06320.173417.19081.2847-52.2863
85.63082.86041.18128.67471.66776.6234-0.0080.1102-0.2481-0.20250.0532-0.24080.3140.2369-0.08180.15910.01750.01520.1965-0.01830.180927.2595-8.3448-43.9612
98.4134-2.4921-2.75152.2978-0.05288.2161-0.01270.1879-0.6298-0.1475-0.1307-0.01820.7530.61210.16270.36030.0157-0.00880.2609-0.00460.208820.69598.9861-13.5283
108.6814-4.8713-4.19555.96122.87424.38370.03170.17140.4694-0.30570.0819-0.3784-0.43380.1683-0.19340.3361-0.048-0.01220.20730.03970.237821.383322.0917-14.3373
113.2591-1.97413.02524.4753-0.10014.77320.1730.28530.1369-0.2695-0.06520.0734-0.404-0.0628-0.13680.3761-0.00170.06440.26880.04330.205818.386916.4255-23.2007
126.31390.5851-2.41672.49980.78582.48690.3430.10070.3771-0.9588-0.50860.6161-0.6714-0.77850.08630.53040.1973-0.08280.5926-0.01840.31571.770218.3451-28.7905
139.111-4.30262.25275.9501-1.14044.8966-0.04510.0428-0.1535-0.11450.05830.3903-0.0401-0.7773-0.04510.343-0.00830.0050.3826-0.00990.14339.168.5273-22.2112
144.22620.26-1.10652.0832-0.80033.21420.04420.138-0.03840.01160.00860.05510.0013-0.1995-0.05460.2912-0.0189-0.0150.23990.00260.135211.885411.1817-11.7755
159.3115-0.9706-0.1124.055-0.87154.52670.0282-0.02120.5790.37580.00570.0701-0.5665-0.2120.02410.35610.00320.02930.2057-0.04110.14677.349816.2649-3.4273
166.70193.7344-2.6547.09781.11396.9710.1676-0.39460.26680.2525-0.45220.0535-0.7986-0.31390.20380.4112-0.0046-0.04130.2604-0.04440.292217.785724.1136-1.5979
174.6605-1.99320.22069.41040.96335.8054-0.1407-0.4070.17570.49120.3923-0.6254-0.07780.4325-0.23120.2507-0.0061-0.05630.2654-0.03650.238826.200615.4568-5.7304
186.1382-0.79680.67052.58280.62013.9334-0.03380.57760.3786-0.8381-0.2357-0.15570.09770.55210.23390.33880.02350.03520.31120.04030.401933.450850.3891-15.1726
195.93712.0877-3.97744.1347-1.52875.883-0.32750.062-0.6639-0.0294-0.0156-0.47750.3510.0190.38550.28280.00270.03710.21630.02490.411332.903138.3967-8.7686
203.83281.96562.29273.63591.93522.11960.0761-0.1056-0.45050.1350.0175-0.20680.19030.1612-0.03050.19340.02670.0330.2720.0660.392241.272745.4156-5.698
217.8549-2.4691-7.73813.1584.238.99310.2024-1.1554-0.83711.4447-0.2396-0.31290.06720.4651-0.02110.5072-0.1599-0.07340.57480.20840.354341.841249.801310.7648
224.3743-0.1004-0.22554.0393-1.37668.3772-0.0239-0.46360.2470.369-0.21460.0995-0.4661-0.15420.24720.1781-0.0342-0.02320.31130.02630.395837.506356.0172-1.0001
234.7764-0.9842-0.96971.95580.62223.4536-0.017-0.17820.04740.0603-0.07040.08140.0295-0.06250.05820.147-0.0135-0.02690.1630.01480.264825.799550.5743-4.2504
245.82833.8667-2.52787.4671-3.01527.3082-0.38410.4804-0.4227-0.44010.17930.16930.6837-0.43940.12480.2219-0.0280.00190.2468-0.04270.295724.29740.2621-15.1394
252.87291.88212.66876.31531.62152.765-0.10090.3537-0.6663-0.63120.198-0.17850.1061-0.01050.05380.295-0.03160.07220.3278-0.07240.379355.328857.1278-14.2577
266.14341.40932.93646.1283.0925.077-0.20960.36560.3639-0.5040.04960.0404-0.44460.00730.11130.25820.0110.01960.15810.02690.184853.781869.9853-13.8018
272.45010.2071-2.2673.44910.01622.22030.09820.00890.06680.0685-0.00930.1081-0.4531-0.5436-0.09730.20210.0118-0.00410.2790.00490.296945.955263.42-6.1189
285.1707-4.14976.95523.4912-5.43489.7533-0.2102-0.92780.39671.74540.0260.2489-0.7685-0.23190.08990.5293-0.00360.03260.5071-0.07640.338146.303163.622310.6042
293.5181-0.4687-0.81295.49053.07421.9912-0.0639-0.467-0.22710.46850.0703-0.1760.4620.1676-0.02020.1762-0.01220.00210.27970.06570.290950.118754.6078-0.9858
305.857-1.33652.10351.305-0.32773.8576-0.051-0.1459-0.1542-0.01640.1262-0.1617-0.0527-0.0157-0.07530.1476-0.00020.0440.1753-0.00540.236459.331458.1615-5.9696
313.0156-0.55271.07042.6991-2.1233.8778-0.06830.09490.49690.19980.0119-0.4491-0.67340.31170.02660.2481-0.07040.00740.2588-0.01340.404967.861367.1523-7.2414
327.00160.4925-1.08277.75592.93946.2186-0.30561.00970.3182-1.0570.5398-0.6985-0.55530.6277-0.17960.3031-0.0940.06950.47120.03640.244161.217264.9534-20.6427
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1chain 'A' and (resid 16 through 25 )A16 - 25
2X-RAY DIFFRACTION2chain 'A' and (resid 26 through 45 )A26 - 45
3X-RAY DIFFRACTION3chain 'A' and (resid 46 through 62 )A46 - 62
4X-RAY DIFFRACTION4chain 'A' and (resid 63 through 69 )A63 - 69
5X-RAY DIFFRACTION5chain 'A' and (resid 70 through 90 )A70 - 90
6X-RAY DIFFRACTION6chain 'A' and (resid 91 through 122 )A91 - 122
7X-RAY DIFFRACTION7chain 'A' and (resid 123 through 162 )A123 - 162
8X-RAY DIFFRACTION8chain 'A' and (resid 163 through 189 )A163 - 189
9X-RAY DIFFRACTION9chain 'B' and (resid 16 through 25 )B16 - 25
10X-RAY DIFFRACTION10chain 'B' and (resid 26 through 45 )B26 - 45
11X-RAY DIFFRACTION11chain 'B' and (resid 46 through 62 )B46 - 62
12X-RAY DIFFRACTION12chain 'B' and (resid 63 through 69 )B63 - 69
13X-RAY DIFFRACTION13chain 'B' and (resid 70 through 90 )B70 - 90
14X-RAY DIFFRACTION14chain 'B' and (resid 91 through 122 )B91 - 122
15X-RAY DIFFRACTION15chain 'B' and (resid 123 through 162 )B123 - 162
16X-RAY DIFFRACTION16chain 'B' and (resid 163 through 171 )B163 - 171
17X-RAY DIFFRACTION17chain 'B' and (resid 172 through 189 )B172 - 189
18X-RAY DIFFRACTION18chain 'C' and (resid 14 through 25 )C14 - 25
19X-RAY DIFFRACTION19chain 'C' and (resid 26 through 45 )C26 - 45
20X-RAY DIFFRACTION20chain 'C' and (resid 46 through 62 )C46 - 62
21X-RAY DIFFRACTION21chain 'C' and (resid 63 through 69 )C63 - 69
22X-RAY DIFFRACTION22chain 'C' and (resid 70 through 90 )C70 - 90
23X-RAY DIFFRACTION23chain 'C' and (resid 91 through 162 )C91 - 162
24X-RAY DIFFRACTION24chain 'C' and (resid 163 through 189 )C163 - 189
25X-RAY DIFFRACTION25chain 'D' and (resid 16 through 25 )D16 - 25
26X-RAY DIFFRACTION26chain 'D' and (resid 26 through 46 )D26 - 46
27X-RAY DIFFRACTION27chain 'D' and (resid 47 through 63 )D47 - 63
28X-RAY DIFFRACTION28chain 'D' and (resid 64 through 69 )D64 - 69
29X-RAY DIFFRACTION29chain 'D' and (resid 70 through 90 )D70 - 90
30X-RAY DIFFRACTION30chain 'D' and (resid 91 through 122 )D91 - 122
31X-RAY DIFFRACTION31chain 'D' and (resid 123 through 171 )D123 - 171
32X-RAY DIFFRACTION32chain 'D' and (resid 172 through 189 )D172 - 189

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