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- PDB-8a0f: Crystal structure of human deoxyhypusine synthase variant K329A i... -

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Basic information

Entry
Database: PDB / ID: 8a0f
TitleCrystal structure of human deoxyhypusine synthase variant K329A in complex with NAD and SPD
ComponentsDeoxyhypusine synthase
KeywordsTRANSFERASE / Hypusination / posttranslational modification
Function / homology
Function and homology information


deoxyhypusine synthase / Hypusine synthesis from eIF5A-lysine / peptidyl-lysine modification to peptidyl-hypusine / deoxyhypusine synthase activity / spermidine metabolic process / spermidine catabolic process / positive regulation of T cell proliferation / glucose homeostasis / translation / positive regulation of cell population proliferation ...deoxyhypusine synthase / Hypusine synthesis from eIF5A-lysine / peptidyl-lysine modification to peptidyl-hypusine / deoxyhypusine synthase activity / spermidine metabolic process / spermidine catabolic process / positive regulation of T cell proliferation / glucose homeostasis / translation / positive regulation of cell population proliferation / identical protein binding / cytoplasm / cytosol
Similarity search - Function
Deoxyhypusine synthase / Deoxyhypusine synthase superfamily / Deoxyhypusine synthase / DHS-like NAD/FAD-binding domain superfamily
Similarity search - Domain/homology
NICOTINAMIDE-ADENINE-DINUCLEOTIDE / SPERMIDINE / Deoxyhypusine synthase
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 1.64 Å
AuthorsWator, E. / Wilk, P. / Grudnik, P.
Funding support Poland, 2items
OrganizationGrant numberCountry
Polish National Science CentreUMO-2019/33/B/NZ1/01839 Poland
Foundation for Polish ScienceTEAM TECH CORE FACILITY/2017-4/6 Poland
CitationJournal: Nat Commun / Year: 2023
Title: Cryo-EM structure of human eIF5A-DHS complex reveals the molecular basis of hypusination-associated neurodegenerative disorders.
Authors: Elżbieta Wątor / Piotr Wilk / Artur Biela / Michał Rawski / Krzysztof M Zak / Wieland Steinchen / Gert Bange / Sebastian Glatt / Przemysław Grudnik /
Abstract: Hypusination is a unique post-translational modification of the eukaryotic translation factor 5A (eIF5A) that is essential for overcoming ribosome stalling at polyproline sequence stretches. The ...Hypusination is a unique post-translational modification of the eukaryotic translation factor 5A (eIF5A) that is essential for overcoming ribosome stalling at polyproline sequence stretches. The initial step of hypusination, the formation of deoxyhypusine, is catalyzed by deoxyhypusine synthase (DHS), however, the molecular details of the DHS-mediated reaction remained elusive. Recently, patient-derived variants of DHS and eIF5A have been linked to rare neurodevelopmental disorders. Here, we present the cryo-EM structure of the human eIF5A-DHS complex at 2.8 Å resolution and a crystal structure of DHS trapped in the key reaction transition state. Furthermore, we show that disease-associated DHS variants influence the complex formation and hypusination efficiency. Hence, our work dissects the molecular details of the deoxyhypusine synthesis reaction and reveals how clinically-relevant mutations affect this crucial cellular process.
History
DepositionMay 27, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 5, 2023Provider: repository / Type: Initial release
Revision 1.1Feb 7, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Deoxyhypusine synthase
B: Deoxyhypusine synthase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)84,1238
Polymers82,3252
Non-polymers1,7986
Water7,981443
1
A: Deoxyhypusine synthase
B: Deoxyhypusine synthase
hetero molecules

A: Deoxyhypusine synthase
B: Deoxyhypusine synthase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)168,24616
Polymers164,6504
Non-polymers3,59512
Water724
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation4_555y,x,-z1
Buried area29740 Å2
ΔGint-110 kcal/mol
Surface area38960 Å2
MethodPISA
Unit cell
Length a, b, c (Å)105.241, 105.241, 160.220
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number154
Space group name H-MP3221
Symmetry operation#1: x,y,z
#2: -y,x-y,z+2/3
#3: -x+y,-x,z+1/3
#4: x-y,-y,-z+1/3
#5: -x,-x+y,-z+2/3
#6: y,x,-z
Components on special symmetry positions
IDModelComponents
11B-679-

HOH

21B-700-

HOH

Noncrystallographic symmetry (NCS)NCS domain:
IDEns-IDDetails
11(chain A and (resid 28 through 31 or resid 33...
21(chain B and (resid 28 through 31 or resid 33...

NCS domain segments:
Dom-IDComponent-IDEns-IDSelection detailsAuth asym-IDAuth seq-ID
111(chain A and (resid 28 through 31 or resid 33...A28 - 31
121(chain A and (resid 28 through 31 or resid 33...A33 - 42
131(chain A and (resid 28 through 31 or resid 33...A44
141(chain A and (resid 28 through 31 or resid 33...A48
151(chain A and (resid 28 through 31 or resid 33...A50 - 60
161(chain A and (resid 28 through 31 or resid 33...A62 - 70
171(chain A and (resid 28 through 31 or resid 33...A72 - 776
181(chain A and (resid 28 through 31 or resid 33...A138 - 155
191(chain A and (resid 28 through 31 or resid 33...A1588
1101(chain A and (resid 28 through 31 or resid 33...A190 - 191
1111(chain A and (resid 28 through 31 or resid 33...A193 - 263
1121(chain A and (resid 28 through 31 or resid 33...A265 - 279
1131(chain A and (resid 28 through 31 or resid 33...A281 - 285
1141(chain A and (resid 28 through 31 or resid 33...A287 - 326
1151(chain A and (resid 28 through 31 or resid 33...A328 - 332
1161(chain A and (resid 28 through 31 or resid 33...A334 - 345
1171(chain A and (resid 28 through 31 or resid 33...A347 - 356
1181(chain A and (resid 28 through 31 or resid 33...A358 - 359
1191(chain A and (resid 28 through 31 or resid 33...A361 - 401
211(chain B and (resid 28 through 31 or resid 33...B28 - 31
221(chain B and (resid 28 through 31 or resid 33...B33 - 42
231(chain B and (resid 28 through 31 or resid 33...B44
241(chain B and (resid 28 through 31 or resid 33...B48
251(chain B and (resid 28 through 31 or resid 33...B50 - 60
261(chain B and (resid 28 through 31 or resid 33...B62 - 70
271(chain B and (resid 28 through 31 or resid 33...B72 - 77
281(chain B and (resid 28 through 31 or resid 33...B92 - 136
291(chain B and (resid 28 through 31 or resid 33...B138 - 153
2101(chain B and (resid 28 through 31 or resid 33...B155 - 156
2111(chain B and (resid 28 through 31 or resid 33...B158 - 188
2121(chain B and (resid 28 through 31 or resid 33...B190 - 191
2131(chain B and (resid 28 through 31 or resid 33...B193 - 263
2141(chain B and (resid 28 through 31 or resid 33...B265 - 279
2151(chain B and (resid 28 through 31 or resid 33...B281 - 285
2161(chain B and (resid 28 through 31 or resid 33...B287 - 326
2171(chain B and (resid 28 through 31 or resid 33...B328 - 332
2181(chain B and (resid 28 through 31 or resid 33...B334 - 345
2191(chain B and (resid 28 through 31 or resid 33...B347 - 356
2201(chain B and (resid 28 through 31 or resid 33...B358 - 359
2211(chain B and (resid 28 through 31 or resid 33...B361 - 401

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Components

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Protein , 1 types, 2 molecules AB

#1: Protein Deoxyhypusine synthase / DHS


Mass: 41162.594 Da / Num. of mol.: 2 / Mutation: K329A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DHPS, DS / Production host: Escherichia coli (E. coli) / References: UniProt: P49366, deoxyhypusine synthase

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Non-polymers , 5 types, 449 molecules

#2: Chemical ChemComp-SPD / SPERMIDINE / N-(2-AMINO-PROPYL)-1,4-DIAMINOBUTANE / PA(34)


Mass: 145.246 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C7H19N3
#3: Chemical ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL


Mass: 62.068 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C2H6O2
#4: Chemical ChemComp-NAD / NICOTINAMIDE-ADENINE-DINUCLEOTIDE


Mass: 663.425 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C21H27N7O14P2 / Comment: NAD*YM
#5: Chemical ChemComp-MRD / (4R)-2-METHYLPENTANE-2,4-DIOL


Mass: 118.174 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H14O2 / Comment: precipitant*YM
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 443 / 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: 3.11 Å3/Da / Density % sol: 60.5 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 8.5
Details: Protein (20 mg/ml) mixed with with equal volume of mother liquor: 0.025-0.125 mM carboxylic acid mix, 30-60% precipitant mix (MPD, PEG 1000, PEG 3350) and 100mM Tris-Bicine pH 8.5. ...Details: Protein (20 mg/ml) mixed with with equal volume of mother liquor: 0.025-0.125 mM carboxylic acid mix, 30-60% precipitant mix (MPD, PEG 1000, PEG 3350) and 100mM Tris-Bicine pH 8.5. conditions based on Morpheus screen.

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: BESSY / Beamline: 14.1 / Wavelength: 0.9184 Å
DetectorType: DECTRIS PILATUS3 S 6M / Detector: PIXEL / Date: Oct 23, 2021
RadiationMonochromator: DCM Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9184 Å / Relative weight: 1
ReflectionResolution: 1.64→46.08 Å / Num. obs: 124222 / % possible obs: 99.1 % / Redundancy: 11.119 % / Biso Wilson estimate: 25.75 Å2 / CC1/2: 0.999 / Rmerge(I) obs: 0.137 / Rrim(I) all: 0.144 / Χ2: 0.78 / Net I/σ(I): 11.5 / Num. measured all: 1381279
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
1.64-1.7411.2263.2030.7121302420047189760.4533.35594.7
1.74-1.8611.2821.7781.3321322618900189000.7251.862100
1.86-2.0110.7890.8842.6518942417560175570.8890.928100
2.01-2.211.5640.4675.2318803816261162600.9660.489100
2.2-2.4611.1570.2499.3816406514707147050.9890.261100
2.46-2.8411.3220.14615.7714772113048130470.9950.153100
2.84-3.4811.0940.08826.7612305311092110920.9980.092100
3.48-4.910.5040.05342.8491152867986780.9990.056100
4.9-46.0810.3010.0449.2351576501850070.9990.04299.8

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Processing

Software
NameVersionClassification
PHENIX1.20.1_4487refinement
XSCALEdata scaling
PDB_EXTRACT3.27data extraction
PHASERphasing
XDSdata reduction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 6XXJ

6xxj


Resolution: 1.64→46.08 Å / SU ML: 0.23 / Cross valid method: THROUGHOUT / σ(F): 1.33 / Phase error: 20.28 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.1777 2095 1.69 %
Rwork0.1559 122004 -
obs0.1563 124099 99.04 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.1 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 140.58 Å2 / Biso mean: 34.3239 Å2 / Biso min: 17.21 Å2
Refinement stepCycle: final / Resolution: 1.64→46.08 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms5113 0 120 443 5676
Biso mean--32.03 44.11 -
Num. residues----653
Refine LS restraints NCS
Ens-IDDom-IDAuth asym-IDNumberRefine-IDRmsType
11A1698X-RAY DIFFRACTION1.111TORSIONAL
12B1698X-RAY DIFFRACTION1.111TORSIONAL
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 15

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
1.64-1.680.42491190.39557008712786
1.68-1.720.38071400.329981438283100
1.72-1.770.31291390.300980918230100
1.77-1.820.30661400.261681698309100
1.82-1.880.26861410.227681708311100
1.88-1.950.22751390.204881048243100
1.95-2.030.21361400.185781758315100
2.03-2.120.17731400.149680998239100
2.12-2.230.18921410.143382298370100
2.23-2.370.14431400.13481478287100
2.37-2.550.14921420.128382328374100
2.55-2.810.15721420.128982378379100
2.81-3.210.1521420.132582748416100
3.21-4.050.14781440.131383458489100
4.05-46.080.16781460.149885818727100
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)
10.05610.03480.03910.0242-0.00690.05420.10870.10730.1892-0.2159-0.0338-0.0903-0.4824-0.12770.00070.44060.09560.01870.2159-0.00280.3111-38.5217-15.8095-6.7534
20.27530.0552-0.06790.0136-0.0160.0160.1152-0.11060.1965-0.1384-0.1919-0.1144-0.26980.1044-0.07320.42650.03360.0480.11640.01580.3058-24.9871-17.4816-15.113
30.356-0.14260.19820.37540.27970.46620.16170.19260.0107-0.3618-0.1556-0.0227-0.1452-0.0055-0.0210.41510.10650.03230.21340.03490.2259-22.2075-30.9237-28.6665
40.1575-0.02190.05010.04170.02160.06860.1495-0.0416-0.2004-0.0468-0.08260.11280.3289-0.11130.00050.32040.0168-0.0580.1977-0.00140.3068-31.0878-53.0448-12.7377
50.1753-0.06040.14930.22070.10780.20860.14160.0604-0.193-0.2666-0.20990.08350.10430.15030.1380.44790.141-0.03520.2434-0.03090.2527-21.3538-52.2974-33.7024
60.6261-0.2537-0.00680.5734-0.02180.57890.14490.10090.0857-0.214-0.1307-0.079-0.14410.1223-0.00060.3530.04960.04350.21450.03050.2389-14.0919-35.0699-25.813
70.50810.0546-0.11290.25940.06090.1920.18480.02630.1945-0.0904-0.14630.0324-0.2364-0.04860.00220.37970.05360.03630.15140.02810.2748-25.2421-22.0069-18.3023
80.1042-0.14010.1240.1938-0.02020.17340.0327-0.22-0.08290.00910.02890.05020.2090.1392-0.00540.17660.0158-0.01680.32310.03950.2454-8.7773-53.689112.2622
90.15550.02370.04530.10260.00660.42870.12040.01480.0198-0.1296-0.0848-0.08780.08620.43540.00440.23350.04990.02190.34210.01920.26531.7387-49.4846-8.9773
100.1188-0.05790.09540.0372-0.02560.11810.1242-0.07990.2994-0.0294-0.098-0.237-0.42720.3721-0.00430.3564-0.13650.08580.32760.00220.4198-0.069-22.6603-5.7875
110.6162-0.19840.30820.3629-0.40730.42560.14050.22120.0898-0.225-0.1757-0.1325-0.02240.53460.05950.29780.01590.13630.48380.0660.29248.7309-39.0029-23.7571
120.48-0.10960.09010.4495-0.08890.7170.1223-0.0114-0.0214-0.1219-0.07710.0170.11670.1501-00.230.05090.00930.22340.0120.2265-6.3261-49.6956-11.8903
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1chain 'A' and (resid 28 through 53 )A28 - 53
2X-RAY DIFFRACTION2chain 'A' and (resid 54 through 73 )A54 - 73
3X-RAY DIFFRACTION3chain 'A' and (resid 74 through 142 )A74 - 142
4X-RAY DIFFRACTION4chain 'A' and (resid 143 through 168 )A143 - 168
5X-RAY DIFFRACTION5chain 'A' and (resid 169 through 197 )A169 - 197
6X-RAY DIFFRACTION6chain 'A' and (resid 198 through 327 )A198 - 327
7X-RAY DIFFRACTION7chain 'A' and (resid 328 through 363 )A328 - 363
8X-RAY DIFFRACTION8chain 'B' and (resid 28 through 74 )B28 - 74
9X-RAY DIFFRACTION9chain 'B' and (resid 75 through 142 )B75 - 142
10X-RAY DIFFRACTION10chain 'B' and (resid 143 through 168 )B143 - 168
11X-RAY DIFFRACTION11chain 'B' and (resid 169 through 214 )B169 - 214
12X-RAY DIFFRACTION12chain 'B' and (resid 215 through 363 )B215 - 363

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