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- PDB-8bb1: T3 SAM lyase in complex with S-adenosylmethionine synthase -

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Basic information

Entry
Database: PDB / ID: 8bb1
TitleT3 SAM lyase in complex with S-adenosylmethionine synthase
Components
  • S-Adenosylmethionine lyase
  • S-adenosylmethionine synthase
KeywordsLYASE / SAM lyase / complex
Function / homology
Function and homology information


S-adenosyl-L-methionine lyase / S-adenosyl-L-methionine lyase activity / S-adenosylmethionine cycle / methionine adenosyltransferase / methionine adenosyltransferase activity / S-adenosylmethionine biosynthetic process / potassium ion binding / one-carbon metabolic process / lyase activity / magnesium ion binding ...S-adenosyl-L-methionine lyase / S-adenosyl-L-methionine lyase activity / S-adenosylmethionine cycle / methionine adenosyltransferase / methionine adenosyltransferase activity / S-adenosylmethionine biosynthetic process / potassium ion binding / one-carbon metabolic process / lyase activity / magnesium ion binding / ATP binding / identical protein binding / cytosol
Similarity search - Function
S-adenosyl-L-methionine hydrolase / S-adenosylmethionine synthetase / S-adenosylmethionine synthetase, N-terminal / S-adenosylmethionine synthetase, central domain / S-adenosylmethionine synthetase, C-terminal / S-adenosylmethionine synthetase, conserved site / S-adenosylmethionine synthetase superfamily / S-adenosylmethionine synthetase, N-terminal domain / S-adenosylmethionine synthetase, central domain / S-adenosylmethionine synthetase, C-terminal domain ...S-adenosyl-L-methionine hydrolase / S-adenosylmethionine synthetase / S-adenosylmethionine synthetase, N-terminal / S-adenosylmethionine synthetase, central domain / S-adenosylmethionine synthetase, C-terminal / S-adenosylmethionine synthetase, conserved site / S-adenosylmethionine synthetase superfamily / S-adenosylmethionine synthetase, N-terminal domain / S-adenosylmethionine synthetase, central domain / S-adenosylmethionine synthetase, C-terminal domain / S-adenosylmethionine synthase signature 1. / S-adenosylmethionine synthase signature 2.
Similarity search - Domain/homology
S-Adenosylmethionine lyase / S-adenosylmethionine synthase
Similarity search - Component
Biological speciesEnterobacteria phage T3 (virus)
Escherichia coli (E. coli)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsTriguis, S. / Selmer, M.
Funding support Sweden, 2items
OrganizationGrant numberCountry
Swedish Research Council2017-03827 Sweden
Knut and Alice Wallenberg FoundationEvolution of new genes and proteins Sweden
CitationJournal: Cell Rep / Year: 2023
Title: Phage T3 overcomes the BREX defense through SAM cleavage and inhibition of SAM synthesis by SAM lyase.
Authors: Aleksandr Andriianov / Silvia Trigüis / Alena Drobiazko / Nicolas Sierro / Nikolai V Ivanov / Maria Selmer / Konstantin Severinov / Artem Isaev /
Abstract: Bacteriophage T3 encodes a SAMase that, through cleavage of S-adenosyl methionine (SAM), circumvents the SAM-dependent type I restriction-modification (R-M) defense. We show that SAMase also allows ...Bacteriophage T3 encodes a SAMase that, through cleavage of S-adenosyl methionine (SAM), circumvents the SAM-dependent type I restriction-modification (R-M) defense. We show that SAMase also allows T3 to evade the BREX defense. Although SAM depletion weakly affects BREX methylation, it completely inhibits the defensive function of BREX, suggesting that SAM could be a co-factor for BREX-mediated exclusion of phage DNA, similar to its anti-defense role in type I R-M. The anti-BREX activity of T3 SAMase is mediated not just by enzymatic degradation of SAM but also by direct inhibition of MetK, the host SAM synthase. We present a 2.8 Å cryoelectron microscopy (cryo-EM) structure of the eight-subunit T3 SAMase-MetK complex. Structure-guided mutagenesis reveals that this interaction stabilizes T3 SAMase in vivo, further stimulating its anti-BREX activity. This work provides insights in the versatility of bacteriophage counterdefense mechanisms and highlights the role of SAM as a co-factor of diverse bacterial immunity systems.
History
DepositionOct 12, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 23, 2023Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: S-adenosylmethionine synthase
B: S-adenosylmethionine synthase
C: S-adenosylmethionine synthase
D: S-adenosylmethionine synthase
E: S-Adenosylmethionine lyase
F: S-Adenosylmethionine lyase
G: S-Adenosylmethionine lyase
H: S-Adenosylmethionine lyase
hetero molecules


Theoretical massNumber of molelcules
Total (without water)239,58912
Polymers239,4478
Non-polymers1424
Water43224
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: SAXS, gel filtration, Gel filtration and mass photometry confirms the stoichiometry of the complex.
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
S-adenosylmethionine synthase / AdoMet synthase / MAT / Methionine adenosyltransferase


Mass: 41998.508 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Source: (natural) Escherichia coli (E. coli) / Strain: K12 / References: UniProt: P0A817, methionine adenosyltransferase
#2: Protein
S-Adenosylmethionine lyase / ADOMetase / Adenosylmethionine lyase / SAMase


Mass: 17863.264 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Enterobacteria phage T3 (virus) / Production host: Escherichia coli (E. coli) / Strain (production host): Top10 / References: UniProt: P07693, EC: 2.5.1.4
#3: Chemical
ChemComp-CL / CHLORIDE ION / Chloride


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


Mass: 18.015 Da / Num. of mol.: 24 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeDetailsEntity IDParent-IDSource
1T3 SAM lyase in complex with E. coli S-adenosylmethionine synthase.COMPLEXT3 SAM lyase was recombinantly expressed in E. coli Top10 and co-purified in complex with S-adenosylmethionine synthase from the host.#1-#20RECOMBINANT
2S-adenosylmethionine synthase in complex with T3 SAM lyase.COMPLEXT3 SAM lyase was recombinantly expressed in E. coli Top10 and co-purified in complex with S-adenosylmethionine synthase from the host.#1-#21NATURAL
3T3 SAM lyase in complex with S-adenosylmethionine synthase.COMPLEXT3 SAM lyase was recombinantly expressed in E. coli Top10 and co-purified in complex with S-adenosylmethionine synthase from the host.#1-#21RECOMBINANT
Molecular weight
IDEntity assembly-IDValue (°)Experimental value
110.239 MDaYES
210.239 MDaYES
310.239 MDaYES
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
21Enterobacteria phage T3 (virus)10759
32Escherichia coli (E. coli)562Top10
43Enterobacteria phage T3 (virus)10759
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
21Escherichia coli (E. coli)562Top10
32Escherichia coli (E. coli)562Top10
43Escherichia coli (E. coli)562Top10
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
125 mMTris(Hydroxymethyl)aminomethaneNH2C(CH2OH)31
2150 mMSodium chlorideNaClSodium chloride1
32.5 mM2-MercaptoethanolHSCH2CH2OH1
SpecimenConc.: 0.125 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: Blotting for 3 seconds before plunging.

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 600 nm / Calibrated defocus min: 350 nm / Calibrated defocus max: 2100 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3 sec. / Electron dose: 47.75 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 5447
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV
Image scansSampling size: 5 µm / Width: 5760 / Height: 4092

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Processing

EM software
IDNameVersionCategory
1cryoSPARC3particle selection
2EPU2.11image acquisition
4cryoSPARC3CTF correction
7Coot0.9.3model fitting
9PHENIX1.20.1-4487model refinement
10cryoSPARC3initial Euler assignment
11cryoSPARC3final Euler assignment
12cryoSPARC3classification
13cryoSPARC33D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1445186
SymmetryPoint symmetry: D2 (2x2 fold dihedral)
3D reconstructionResolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 244912 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Atomic model buildingPDB-ID: 1P7L

1p7l

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