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- EMDB-26567: Structure of Human SAMHD1 with Non-Hydrolysable dGTP Analog -

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Basic information

Entry
Database: EMDB / ID: EMD-26567
TitleStructure of Human SAMHD1 with Non-Hydrolysable dGTP Analog
Map dataHuman SAMHD1 with dGTP(alpha)S Nucleotide Final CryoEM Map
Sample
  • Complex: Human SAMHD1 with Non-Hydrolysable dGTP Analog
    • Protein or peptide: Deoxynucleoside triphosphate triphosphohydrolase SAMHD1
  • Ligand: 2'-deoxyguanosine-5'-O-(1-thiotriphosphate)
KeywordsdNTP Triphosphatase / HYDROLASE
Function / homology
Function and homology information


Nucleotide catabolism / Hydrolases; Acting on ester bonds; Triphosphoric-monoester hydrolases / deoxynucleoside triphosphate hydrolase activity / dGTP binding / triphosphoric monoester hydrolase activity / dATP catabolic process / dGTPase activity / tetraspanin-enriched microdomain / dGTP catabolic process / DNA strand resection involved in replication fork processing ...Nucleotide catabolism / Hydrolases; Acting on ester bonds; Triphosphoric-monoester hydrolases / deoxynucleoside triphosphate hydrolase activity / dGTP binding / triphosphoric monoester hydrolase activity / dATP catabolic process / dGTPase activity / tetraspanin-enriched microdomain / dGTP catabolic process / DNA strand resection involved in replication fork processing / deoxyribonucleotide catabolic process / negative regulation of type I interferon-mediated signaling pathway / regulation of innate immune response / somatic hypermutation of immunoglobulin genes / RNA nuclease activity / double-strand break repair via homologous recombination / Interferon alpha/beta signaling / single-stranded DNA binding / site of double-strand break / protein homotetramerization / defense response to virus / nucleic acid binding / immune response / innate immune response / DNA damage response / GTP binding / RNA binding / zinc ion binding / nucleoplasm / identical protein binding / nucleus / plasma membrane
Similarity search - Function
HD domain profile. / HD domain / HD domain / SAM domain (Sterile alpha motif) / SAM domain profile. / Metal dependent phosphohydrolases with conserved 'HD' motif. / Sterile alpha motif. / HD/PDEase domain / Sterile alpha motif domain / Sterile alpha motif/pointed domain superfamily
Similarity search - Domain/homology
Deoxynucleoside triphosphate triphosphohydrolase SAMHD1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.89 Å
AuthorsHuynh KW / Ammirati M / Han S
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nucleic Acids Res / Year: 2022
Title: Phosphorylation of SAMHD1 Thr592 increases C-terminal domain dynamics, tetramer dissociation and ssDNA binding kinetics.
Authors: Benjamin Orris / Kevin W Huynh / Mark Ammirati / Seungil Han / Ben Bolaños / Jason Carmody / Matthew D Petroski / Benedikt Bosbach / David J Shields / James T Stivers /
Abstract: SAM and HD domain containing deoxynucleoside triphosphate triphosphohydrolase 1 (SAMHD1) is driven into its activated tetramer form by binding of GTP activator and dNTP activators/substrates. In ...SAM and HD domain containing deoxynucleoside triphosphate triphosphohydrolase 1 (SAMHD1) is driven into its activated tetramer form by binding of GTP activator and dNTP activators/substrates. In addition, the inactive monomeric and dimeric forms of the enzyme bind to single-stranded (ss) nucleic acids. During DNA replication SAMHD1 can be phosphorylated by CDK1 and CDK2 at its C-terminal threonine 592 (pSAMHD1), localizing the enzyme to stalled replication forks (RFs) to promote their restart. Although phosphorylation has only a small effect on the dNTPase activity and ssDNA binding affinity of SAMHD1, perturbation of the native T592 by phosphorylation decreased the thermal stability of tetrameric SAMHD1 and accelerated tetramer dissociation in the absence and presence of ssDNA (∼15-fold). In addition, we found that ssDNA binds competitively with GTP to the A1 site. A full-length SAMHD1 cryo-EM structure revealed substantial dynamics in the C-terminal domain (which contains T592), which could be modulated by phosphorylation. We propose that T592 phosphorylation increases tetramer dynamics and allows invasion of ssDNA into the A1 site and the previously characterized DNA binding surface at the dimer-dimer interface. These features are consistent with rapid and regiospecific inactivation of pSAMHD1 dNTPase at RFs or other sites of free ssDNA in cells.
History
DepositionMar 31, 2022-
Header (metadata) releaseJul 20, 2022-
Map releaseJul 20, 2022-
UpdateFeb 14, 2024-
Current statusFeb 14, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26567.png

Downloads & links

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Map

FileDownload / File: emd_26567.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHuman SAMHD1 with dGTP(alpha)S Nucleotide Final CryoEM Map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 300 pix.
= 252. Å		0.84 Å/pix.
x 300 pix.
= 252. Å		0.84 Å/pix.
x 300 pix.
= 252. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.84 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.025
Minimum - Maximum-0.08315243 - 0.14433534
Average (Standard dev.)-0.000013746111 (±0.0041794605)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 251.99998 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Human SAMHD1 with dGTP(alpha)S Nucleotide Half Map 2

Fileemd_26567_half_map_1.map
AnnotationHuman SAMHD1 with dGTP(alpha)S Nucleotide Half Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Human SAMHD1 with dGTP(alpha)S Nucleotide Half Map 1

Fileemd_26567_half_map_2.map
AnnotationHuman SAMHD1 with dGTP(alpha)S Nucleotide Half Map 1
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 SAMHD1 with Non-Hydrolysable dGTP Analog

EntireName: Human SAMHD1 with Non-Hydrolysable dGTP Analog
Components
  • Complex: Human SAMHD1 with Non-Hydrolysable dGTP Analog
    • Protein or peptide: Deoxynucleoside triphosphate triphosphohydrolase SAMHD1
  • Ligand: 2'-deoxyguanosine-5'-O-(1-thiotriphosphate)

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Supramolecule #1: Human SAMHD1 with Non-Hydrolysable dGTP Analog

SupramoleculeName: Human SAMHD1 with Non-Hydrolysable dGTP Analog / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 288 KDa

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Macromolecule #1: Deoxynucleoside triphosphate triphosphohydrolase SAMHD1

MacromoleculeName: Deoxynucleoside triphosphate triphosphohydrolase SAMHD1
type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
EC number: Hydrolases; Acting on ester bonds; Triphosphoric-monoester hydrolases
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 72.305414 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MQRADSEQPS KRPRCDDSPR TPSNTPSAEA DWSPGLELHP DYKTWGPEQV CSFLRRGGFE EPVLLKNIRE NEITGALLPC LDESRFENL GVSSLGERKK LLSYIQRLVQ IHVDTMKVIN DPIHGHIELH PLLVRIIDTP QFQRLRYIKQ LGGGYYVFPG A SHNRFEHS ...String:
MQRADSEQPS KRPRCDDSPR TPSNTPSAEA DWSPGLELHP DYKTWGPEQV CSFLRRGGFE EPVLLKNIRE NEITGALLPC LDESRFENL GVSSLGERKK LLSYIQRLVQ IHVDTMKVIN DPIHGHIELH PLLVRIIDTP QFQRLRYIKQ LGGGYYVFPG A SHNRFEHS LGVGYLAGCL VHALGEKQPE LQISERDVLC VQIAGLCHDL GHGPFSHMFD GRFIPLARPE VKWTHEQGSV MM FEHLINS NGIKPVMEQY GLIPEEDICF IKEQIVGPLE SPVEDSLWPY KGRPENKSFL YEIVSNKRNG IDVDKWDYFA RDC HHLGIQ NNFDYKRFIK FARVCEVDNE LRICARDKEV GNLYDMFHTR NSLHRRAYQH KVGNIIDTMI TDAFLKADDY IEIT GAGGK KYRISTAIDD MEAYTKLTDN IFLEILYSTD PKLKDAREIL KQIEYRNLFK YVGETQPTGQ IKIKREDYES LPKEV ASAK PKVLLDVKLK AEDFIVDVIN MDYGMQEKNP IDHVSFYCKT APNRAIRITK NQVSQLLPEK FAEQLIRVYC KKVDRK SLY AARQYFVQWC ADRNFTKPQD GDVIAPLITP QKKEWNDSTS VQNPTRLREA SKSRVQLFKD DPM

UniProtKB: Deoxynucleoside triphosphate triphosphohydrolase SAMHD1

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Macromolecule #2: 2'-deoxyguanosine-5'-O-(1-thiotriphosphate)

MacromoleculeName: 2'-deoxyguanosine-5'-O-(1-thiotriphosphate) / type: ligand / ID: 2 / Number of copies: 12 / Formula: T8T
Molecular weightTheoretical: 523.247 Da
Chemical component information

ChemComp-T8T:
2'-deoxyguanosine-5'-O-(1-thiotriphosphate)

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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

Concentration0.6 mg/mL
BufferpH: 7.5
Component:
ConcentrationNameFormula
50.0 mMHEPES
300.0 mMPotassium ChlorideKCl
2.0 mMMagnesium ChlorideMgCl2
0.5 mMTCEP
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.2 µm / Nominal defocus min: 0.6 µm
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 1 / Number real images: 3222 / Average electron dose: 57.36 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1030155
Startup modelType of model: OTHER / Details: SGD Algorithm in RELION 3.1.0
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1.0)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.89 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 114078
FSC plot (resolution estimation)

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