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- PDB-7zyt: Crystal structure of the I318T pathogenic variant of the human di... -

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Basic information

Entry
Database: PDB / ID: 7zyt
TitleCrystal structure of the I318T pathogenic variant of the human dihydrolipoamide dehydrogenase
ComponentsDihydrolipoyl dehydrogenase, mitochondrial
KeywordsOXIDOREDUCTASE / lipoamide dehydrogenase / pathogenic mutation / E3 deficiency / alpha-ketoglutarate dehydrogenase complex / 2-oxoglutarate dehydrogenase complex / pyruvate dehydrogenase complex
Function / homology
Function and homology information


acetyltransferase complex / acrosomal matrix / OGDH complex synthesizes succinyl-CoA from 2-OG / OADH complex synthesizes glutaryl-CoA from 2-OA / oxoadipate dehydrogenase complex / Glycine degradation / branched-chain alpha-ketoacid dehydrogenase complex / BCKDH synthesizes BCAA-CoA from KIC, KMVA, KIV / Loss-of-function mutations in DBT cause MSUD2 / Loss-of-function mutations in DLD cause MSUD3/DLDD ...acetyltransferase complex / acrosomal matrix / OGDH complex synthesizes succinyl-CoA from 2-OG / OADH complex synthesizes glutaryl-CoA from 2-OA / oxoadipate dehydrogenase complex / Glycine degradation / branched-chain alpha-ketoacid dehydrogenase complex / BCKDH synthesizes BCAA-CoA from KIC, KMVA, KIV / Loss-of-function mutations in DBT cause MSUD2 / Loss-of-function mutations in DLD cause MSUD3/DLDD / H139Hfs13* PPM1K causes a mild variant of MSUD / PDH complex synthesizes acetyl-CoA from PYR / Branched-chain ketoacid dehydrogenase kinase deficiency / dihydrolipoyl dehydrogenase / dihydrolipoyl dehydrogenase (NADH) activity / Regulation of pyruvate dehydrogenase (PDH) complex / pyruvate decarboxylation to acetyl-CoA / branched-chain amino acid catabolic process / pyruvate dehydrogenase complex / oxoglutarate dehydrogenase complex / Branched-chain amino acid catabolism / 2-oxoglutarate metabolic process / motile cilium / Signaling by Retinoic Acid / pyruvate metabolic process / sperm capacitation / mitochondrial electron transport, NADH to ubiquinone / gastrulation / Mitochondrial protein degradation / regulation of membrane potential / flavin adenine dinucleotide binding / mitochondrial matrix / mitochondrion / proteolysis / nucleus
Similarity search - Function
: / Dihydrolipoamide dehydrogenase / Pyridine nucleotide-disulphide oxidoreductase, class I / Pyridine nucleotide-disulphide oxidoreductase, class I, active site / Pyridine nucleotide-disulphide oxidoreductases class-I active site. / Pyridine nucleotide-disulphide oxidoreductase, dimerisation domain / Pyridine nucleotide-disulphide oxidoreductase, dimerisation domain / FAD/NAD-linked reductase, dimerisation domain superfamily / FAD/NAD(P)-binding domain / Pyridine nucleotide-disulphide oxidoreductase / FAD/NAD(P)-binding domain superfamily
Similarity search - Domain/homology
FLAVIN-ADENINE DINUCLEOTIDE / Dihydrolipoyl dehydrogenase, mitochondrial
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 2.892 Å
AuthorsNemes-Nikodem, E. / Szabo, E. / Vass, K.R. / Lennartz, F. / Nagy, B. / Torocsik, B. / Weiss, M.S. / Adam-Vizi, V. / Ambrus, A.
Funding support Hungary, European Union, 3items
OrganizationGrant numberCountry
Hungarian Academy of SciencesKTIA_13_NAP_III/6 and 2017-1.2.1-NKP-2017-00002 [to A.-V.V.] Hungary
European Union (EU)Horizon 2020 Research and Innovation Programme MX-212-00269 ST [to A.A.]European Union
Hungarian National Research, Development and Innovation OfficeTKP2021-EGA-25 Hungary
CitationJournal: Int J Mol Sci / Year: 2023
Title: Structural and Biochemical Investigation of Selected Pathogenic Mutants of the Human Dihydrolipoamide Dehydrogenase.
Authors: Szabo, E. / Nemes-Nikodem, E. / Vass, K.R. / Zambo, Z. / Zrupko, E. / Torocsik, B. / Ozohanics, O. / Nagy, B. / Ambrus, A.
History
DepositionMay 25, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 14, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 26, 2023Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Feb 7, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model
Revision 1.3Nov 13, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Dihydrolipoyl dehydrogenase, mitochondrial
B: Dihydrolipoyl dehydrogenase, mitochondrial
hetero molecules


Theoretical massNumber of molelcules
Total (without water)107,3027
Polymers105,2502
Non-polymers2,0525
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area11370 Å2
ΔGint-51 kcal/mol
Surface area34830 Å2
MethodPISA
Unit cell
Length a, b, c (Å)188.650, 59.000, 83.230
Angle α, β, γ (deg.)90.000, 101.400, 90.000
Int Tables number5
Space group name H-MC121
Components on special symmetry positions
IDModelComponents
11B-503-

BTB

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Components

#1: Protein Dihydrolipoyl dehydrogenase, mitochondrial / Dihydrolipoamide dehydrogenase / Glycine cleavage system L protein


Mass: 52625.117 Da / Num. of mol.: 2 / Mutation: I318T
Source method: isolated from a genetically manipulated source
Details: Sequence of the Strep-tag with the linker amino acids: MASWSHPQFEKGALEVLFQGPG
Source: (gene. exp.) Homo sapiens (human) / Gene: DLD, GCSL, LAD, PHE3 / Plasmid: pET52b+ / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P09622, dihydrolipoyl dehydrogenase
#2: Chemical ChemComp-FAD / FLAVIN-ADENINE DINUCLEOTIDE


Mass: 785.550 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C27H33N9O15P2 / Feature type: SUBJECT OF INVESTIGATION / Comment: FAD*YM
#3: Chemical ChemComp-BTB / 2-[BIS-(2-HYDROXY-ETHYL)-AMINO]-2-HYDROXYMETHYL-PROPANE-1,3-DIOL / BIS-TRIS BUFFER


Mass: 209.240 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C8H19NO5 / Comment: pH buffer*YM
#4: Chemical ChemComp-EDO / 1,2-ETHANEDIOL / ETHYLENE GLYCOL


Mass: 62.068 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C2H6O2
Has ligand of interestY
Has protein modificationY

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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: 2.16 Å3/Da / Density % sol: 42.98 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 6.7
Details: 0.2 M magnesium chloride, 0.1 M BIS-TRIS (pH 6.7), 29 (v/v)% PEG 3350, cryo-protection: 15 and then 30 (v/v)% ethylene glycol in the reservoir

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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 6M / Detector: PIXEL / Date: Oct 23, 2021
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9184 Å / Relative weight: 1
ReflectionResolution: 2.88→44.14 Å / Num. obs: 20384 / % possible obs: 98.3 % / Redundancy: 6.439 % / Biso Wilson estimate: 87.275 Å2 / CC1/2: 0.998 / Rmerge(I) obs: 0.182 / Rrim(I) all: 0.198 / Χ2: 0.74 / Net I/σ(I): 8.37
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
2.88-3.056.1772.9840.5718568330030060.3543.25391.1
3.05-3.266.4681.8171.0120166313531180.5641.97699.5
3.26-3.526.720.9482.0719462290728960.8481.02899.6
3.52-3.856.8710.4754.218194265826480.9560.51499.6
3.85-4.36.5650.2268.1216098245724520.990.24599.8
4.3-4.966.2340.12813.0313435215621550.9950.14100
4.96-6.066.5710.10815.4511972182318220.9960.11899.9
6.06-8.56.0340.05525.818749145114500.9990.0699.9
8.5-44.1395.5060.03241.9246038588360.9990.03597.4

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Phasing

PhasingMethod: molecular replacement
Phasing MRR rigid body: 0.604
Highest resolutionLowest resolution
Rotation47.65 Å3.46 Å

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Processing

Software
NameVersionClassification
MxCuBEdata collection
XDSdata reduction
XSCALEdata scaling
MOLREP11.7.03; 13.07.2020phasing
PHENIX1.14refinement
PDB_EXTRACT3.27data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 6I4Q

6i4q


Resolution: 2.892→44.14 Å / SU ML: 0.66 / Cross valid method: THROUGHOUT / σ(F): 1.33 / Phase error: 41.83 / Stereochemistry target values: ML
Details: refinement was carried out using NCS torsion-angle restraints
RfactorNum. reflection% reflection
Rfree0.287 1010 5 %
Rwork0.2504 19203 -
obs0.2522 20213 99.18 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 194.76 Å2 / Biso mean: 119.314 Å2 / Biso min: 64.24 Å2
Refinement stepCycle: final / Resolution: 2.892→44.14 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms7012 0 138 0 7150
Biso mean--118.01 --
Num. residues----944
LS refinement shell

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

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection Rwork% reflection obs (%)
2.892-3.04450.53291410.5493268599
3.0445-3.23510.54161430.4577271299
3.2351-3.48480.39251440.36982739100
3.4848-3.83540.36341440.2959272899
3.8354-4.38990.29061440.22592758100
4.3899-5.52910.24271460.20782774100
5.5291-44.140.21221480.1947280798

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