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- EMDB-45831: CryoEM Structure of the human full length NAD Kinase -

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Basic information

Entry
Database: EMDB / ID: EMD-45831
TitleCryoEM Structure of the human full length NAD Kinase
Map data
Sample
  • Complex: human NAD Kinase
    • Protein or peptide: NAD kinase
Keywordscatalyzes the reaction of ATP + NAD+ = ADP + H+ + NADP+ / SIGNALING PROTEIN
Function / homology
Function and homology information


NAD+ kinase / NAD+ kinase activity / NADP biosynthetic process / Nicotinate metabolism / NAD metabolic process / positive regulation of insulin secretion involved in cellular response to glucose stimulus / ATP metabolic process / phosphorylation / ATP binding / metal ion binding / cytosol
Similarity search - Function
ATP-NAD kinase C-terminal domain / NAD kinase / ATP-NAD kinase, PpnK-type, C-terminal / ATP-NAD kinase N-terminal domain / NAD kinase/diacylglycerol kinase-like domain superfamily / Inorganic polyphosphate/ATP-NAD kinase, N-terminal
Similarity search - Domain/homology
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.18 Å
AuthorsLi Y / Chen Z / Labesse G / Hoxhaj G
Funding support United States, France, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Cancer Institute (NIH/NCI)R01GM143236 United States
Welch FoundationI-2067-20210327 United States
Welch FoundationI-2067-20240404 United States
Agence Nationale de la Recherche (ANR)ANR-20-CE18-0025 France
Fondation pour la Recherche Medicale (FRM)EQU202303016265 France
CitationJournal: Sci Adv / Year: 2025
Title: Cryo-EM structure and regulation of human NAD kinase.
Authors: Prakash P Praharaj / Yang Li / Charline Mary / Mona H Soflaee / Kevin Ryu / Dohun Kim / Diem H Tran / Trishna Dey / Harrison J Tom / Halie Rion / Muriel Gelin / Andrew Lemoff / Lauren G ...Authors: Prakash P Praharaj / Yang Li / Charline Mary / Mona H Soflaee / Kevin Ryu / Dohun Kim / Diem H Tran / Trishna Dey / Harrison J Tom / Halie Rion / Muriel Gelin / Andrew Lemoff / Lauren G Zacharias / João S Patricio / Thomas P Mathews / Zhe Chen / Corinne Lionne / Gerta Hoxhaj / Gilles Labesse /
Abstract: Reduced nicotinamide adenine dinucleotide phosphate (NADPH) is a crucial reducing cofactor for reductive biosynthesis and protection from oxidative stress. To fulfill their heightened anabolic and ...Reduced nicotinamide adenine dinucleotide phosphate (NADPH) is a crucial reducing cofactor for reductive biosynthesis and protection from oxidative stress. To fulfill their heightened anabolic and reductive power demands, cancer cells must boost their NADPH production. Progrowth and mitogenic protein kinases promote the activity of cytosolic NAD kinase (NADK), which produces NADP, a limiting NADPH precursor. However, the molecular architecture and mechanistic regulation of human NADK remain undescribed. Here, we report the cryo-electron microscopy structure of human NADK, both in its apo-form and in complex with its substrate NAD (nicotinamide adenine dinucleotide), revealing a tetrameric organization with distinct structural features. We discover that the amino (N)- and carboxyl (C)-terminal tails of NADK have opposing effects on its enzymatic activity and cellular NADP(H) levels. Specifically, the C-terminal region is critical for NADK activity, whereas the N-terminal region exhibits an inhibitory role. This study highlights molecular insights into the regulation of a vital enzyme governing NADP(H) production.
History
DepositionJul 20, 2024-
Header (metadata) releaseJan 8, 2025-
Map releaseJan 8, 2025-
UpdateFeb 12, 2025-
Current statusFeb 12, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45831.png

Downloads & links

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Map

FileDownload / File: emd_45831.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 300 pix.
= 249. Å		0.83 Å/pix.
x 300 pix.
= 249. Å		0.83 Å/pix.
x 300 pix.
= 249. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.2
Minimum - Maximum-0.0017390029 - 1.7437769
Average (Standard dev.)0.0015870417 (±0.030643428)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 249.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_45831_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_45831_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : human NAD Kinase

EntireName: human NAD Kinase
Components
  • Complex: human NAD Kinase
    • Protein or peptide: NAD kinase

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Supramolecule #1: human NAD Kinase

SupramoleculeName: human NAD Kinase / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all / Details: Tetramer of human NAD Kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 49 kDa/nm

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Macromolecule #1: NAD kinase

MacromoleculeName: NAD kinase / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO / EC number: NAD+ kinase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 52.892996 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MEMEQEKMTM NKELSPDAAA YCCSACHGDE TWSYNHPIRG RAKSRSLSAS PALGSTKEFR RTRSLHGPCP VTTFGPKACV LQNPQTIMH IQDPASQRLT WNKSPKSVLV IKKMRDASLL QPFKELCTHL MEENMIVYVE KKVLEDPAIA SDESFGAVKK K FCTFREDY ...String:
MEMEQEKMTM NKELSPDAAA YCCSACHGDE TWSYNHPIRG RAKSRSLSAS PALGSTKEFR RTRSLHGPCP VTTFGPKACV LQNPQTIMH IQDPASQRLT WNKSPKSVLV IKKMRDASLL QPFKELCTHL MEENMIVYVE KKVLEDPAIA SDESFGAVKK K FCTFREDY DDISNQIDFI ICLGGDGTLL YASSLFQGSV PPVMAFHLGS LGFLTPFSFE NFQSQVTQVI EGNAAVVLRS RL KVRVVKE LRGKKTAVHN GLGENGSQAA GLDMDVGKQA MQYQVLNEVV IDRGPSSYLS NVDVYLDGHL ITTVQGDGVI VST PTGSTA YAAAAGASMI HPNVPAIMIT PICPHSLSFR PIVVPAGVEL KIMLSPEARN TAWVSFDGRK RQEIRHGDSI SITT SCYPL PSICVRDPVS DWFESLAQCL HWNVRKKQAH FEEEEEEEEE GTRTRPLEQK LISEEDLAAN DILDYKDDDD KV

UniProtKB: NAD kinase

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.2 mg/mL
BufferpH: 7.4
Component:
ConcentrationName
10.0 mMHEPES
0.01 %NP-40
1.0 mMDTT

Details: 0.2 mg/ml in 10 mM HEPES, 0.01% NP-40, and 1 mM DTT.
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 80 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.038 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV
DetailsPurified by FPLC. Monodispersed.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 6013 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 105000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1820219 / Details: template based picking using a low resolution map
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: D2 (2x2 fold dihedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 3.18 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 77606
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 3 / Software - Name: cryoSPARC
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL
Output model

PDB-9cr3:
CryoEM Structure of the human full length NAD Kinase

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