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- EMDB-13828: Single Particle Cryo-EM structure of photosynthetic A10B10 glycer... -

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Basic information

Entry
Database: EMDB / ID: EMD-13828
TitleSingle Particle Cryo-EM structure of photosynthetic A10B10 glyceraldehyde-3-phospahte dehydrogenase from Spinacia oleracea.
Map dataA10B10 photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) icosamer.
Sample
  • Complex: A10B10 glyceraldehyde-3-phospahte dehydrogenase hetero-icosamer complexed with NAD.
    • Protein or peptide: Glyceraldehyde-3-phosphate dehydrogenase B, chloroplastic
    • Protein or peptide: Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic
  • Ligand: NICOTINAMIDE-ADENINE-DINUCLEOTIDE
KeywordsPhotosynthesis / Calvin-Benson cycle / redox regulation / OXIDOREDUCTASE
Function / homology
Function and homology information


glyceraldehyde-3-phosphate dehydrogenase (NADP+) (phosphorylating) / glyceraldehyde-3-phosphate dehydrogenase (NADP+) (phosphorylating) activity / reductive pentose-phosphate cycle / glyceraldehyde-3-phosphate dehydrogenase (NAD+) (phosphorylating) activity / chloroplast / glucose metabolic process / NAD binding / NADP binding
Similarity search - Function
Glyceraldehyde 3-phosphate dehydrogenase, active site / Glyceraldehyde 3-phosphate dehydrogenase active site. / Glyceraldehyde-3-phosphate dehydrogenase, type I / Glyceraldehyde 3-phosphate dehydrogenase, NAD binding domain / Glyceraldehyde 3-phosphate dehydrogenase, NAD(P) binding domain / Glyceraldehyde 3-phosphate dehydrogenase, catalytic domain / Glyceraldehyde/Erythrose phosphate dehydrogenase family / Glyceraldehyde 3-phosphate dehydrogenase, C-terminal domain / Glyceraldehyde 3-phosphate dehydrogenase, NAD binding domain / NAD(P)-binding domain superfamily
Similarity search - Domain/homology
Glyceraldehyde-3-phosphate dehydrogenase B, chloroplastic / Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic
Similarity search - Component
Biological speciesSpinacia oleracea (spinach)
Methodsingle particle reconstruction / cryo EM / Resolution: 13.0 Å
AuthorsMarotta R / Fermani S
Funding support France, 1 items
OrganizationGrant numberCountry
Grenoble Instruct-ERIC Center (ISBG)PID 1829 France
CitationJournal: Acta Crystallogr D Struct Biol / Year: 2022
Title: Unravelling the regulation pathway of photosynthetic AB-GAPDH.
Authors: Roberto Marotta / Alessandra Del Giudice / Libero Gurrieri / Silvia Fanti / Paolo Swuec / Luciano Galantini / Giuseppe Falini / Paolo Trost / Simona Fermani / Francesca Sparla /
Abstract: Oxygenic phototrophs perform carbon fixation through the Calvin-Benson cycle. Different mechanisms adjust the cycle and the light-harvesting reactions to rapid environmental changes. Photosynthetic ...Oxygenic phototrophs perform carbon fixation through the Calvin-Benson cycle. Different mechanisms adjust the cycle and the light-harvesting reactions to rapid environmental changes. Photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a key enzyme in the cycle. In land plants, different photosynthetic GAPDHs exist: the most abundant isoform is formed by AB heterotetramers and the least abundant by A homotetramers. Regardless of the subunit composition, GAPDH is the major consumer of photosynthetic NADPH and its activity is strictly regulated. While A-GAPDH is regulated by CP12, AB-GAPDH is autonomously regulated through the C-terminal extension (CTE) of its B subunits. Reversible inhibition of AB-GAPDH occurs via the oxidation of a cysteine pair located in the CTE and the substitution of NADP(H) with NAD(H) in the cofactor-binding site. These combined conditions lead to a change in the oligomerization state and enzyme inhibition. SEC-SAXS and single-particle cryo-EM analysis were applied to reveal the structural basis of this regulatory mechanism. Both approaches revealed that spinach (AB)-GAPDH oligomers with n = 1, 2, 4 and 5 co-exist in a dynamic system. B subunits mediate the contacts between adjacent tetramers in AB and AB oligomers. The CTE of each B subunit penetrates into the active site of a B subunit of the adjacent tetramer, which in turn moves its CTE in the opposite direction, effectively preventing the binding of the substrate 1,3-bisphosphoglycerate in the B subunits. The whole mechanism is made possible, and eventually controlled, by pyridine nucleotides. In fact, NAD(H), by removing NADP(H) from A subunits, allows the entrance of the CTE into the active site of the B subunit, hence stabilizing inhibited oligomers.
History
DepositionNov 2, 2021-
Header (metadata) releaseNov 16, 2022-
Map releaseNov 16, 2022-
UpdateJul 17, 2024-
Current statusJul 17, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_13828.png

Downloads & links

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Map

FileDownload / File: emd_13828.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationA10B10 photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) icosamer.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.21 Å/pix.
x 300 pix.
= 363. Å		1.21 Å/pix.
x 300 pix.
= 363. Å		1.21 Å/pix.
x 300 pix.
= 363. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.21 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.013
Minimum - Maximum-0.008460227 - 0.0262652
Average (Standard dev.)0.0003046998 (±0.002465734)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 363.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: A10B10 photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) icosamer....

Fileemd_13828_half_map_1.map
AnnotationA10B10 photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) icosamer.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: A10B10 photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) icosamer....

Fileemd_13828_half_map_2.map
AnnotationA10B10 photosynthetic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) icosamer.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : A10B10 glyceraldehyde-3-phospahte dehydrogenase hetero-icosamer c...

EntireName: A10B10 glyceraldehyde-3-phospahte dehydrogenase hetero-icosamer complexed with NAD.
Components
  • Complex: A10B10 glyceraldehyde-3-phospahte dehydrogenase hetero-icosamer complexed with NAD.
    • Protein or peptide: Glyceraldehyde-3-phosphate dehydrogenase B, chloroplastic
    • Protein or peptide: Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic
  • Ligand: NICOTINAMIDE-ADENINE-DINUCLEOTIDE

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Supramolecule #1: A10B10 glyceraldehyde-3-phospahte dehydrogenase hetero-icosamer c...

SupramoleculeName: A10B10 glyceraldehyde-3-phospahte dehydrogenase hetero-icosamer complexed with NAD.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Spinacia oleracea (spinach)

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Macromolecule #1: Glyceraldehyde-3-phosphate dehydrogenase B, chloroplastic

MacromoleculeName: Glyceraldehyde-3-phosphate dehydrogenase B, chloroplastic
type: protein_or_peptide / ID: 1 / Number of copies: 10 / Enantiomer: LEVO
EC number: glyceraldehyde-3-phosphate dehydrogenase (NADP+) (phosphorylating)
Source (natural)Organism: Spinacia oleracea (spinach)
Molecular weightTheoretical: 48.183789 KDa
SequenceString: MASHAALAPS RIPASTRLAS KASQQYSFLT QCSFKRLDVA DFSGLRSSNS VTFTREASFH DVIAAQLTTK PTGAAPVRGE TVAKLKVAI NGFGRIGRNF LRCWHGRKDS PLDVVVVNDS GGVKSATHLL KYDSILGTFK ADVKIIDNET FSIDGKPIKV V SNRDPLKL ...String:
MASHAALAPS RIPASTRLAS KASQQYSFLT QCSFKRLDVA DFSGLRSSNS VTFTREASFH DVIAAQLTTK PTGAAPVRGE TVAKLKVAI NGFGRIGRNF LRCWHGRKDS PLDVVVVNDS GGVKSATHLL KYDSILGTFK ADVKIIDNET FSIDGKPIKV V SNRDPLKL PWAELGIDIV IEGTGVFVDG PGAGKHIQAG AKKVIITAPA KGSDIPTYVV GVNEKDYGHD VANIISNASC TT NCLAPFV KVLDEELGIV KGTMTTTHSY TGDQRLLDAS HRDLRRARAA ALNIVPTSTG AAKAVSLVLP QLKGKLNGIA LRV PTPNVS VVDLVVNIEK VGVTAEDVNN AFRKAAAGPL KGVLDVCDIP LVSVDFRCSD FSSTIDSSLT MVMGGDMVKV VAWY DNEWG YSQRVVDLAD LVANKWPGLE GSVASGDPLE DFCKDNPADE ECKLYE

UniProtKB: Glyceraldehyde-3-phosphate dehydrogenase B, chloroplastic

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Macromolecule #2: Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glycera...

MacromoleculeName: Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3- ...Name: Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic,Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic
type: protein_or_peptide / ID: 2 / Number of copies: 10 / Enantiomer: LEVO
EC number: glyceraldehyde-3-phosphate dehydrogenase (NADP+) (phosphorylating)
Source (natural)Organism: Spinacia oleracea (spinach)
Molecular weightTheoretical: 36.256391 KDa
SequenceString: KLKVAINGFG RIGRNFLRCW HGRKDSPLDV VVINDTGGVK QASHLLKYDS ILGTFDADVK TAGDSAISVD GKVIKVVSDR NPVNLPWGD MGIDLVIEGT GVFVDRDGAG KHLQAGAKKV LITAPGKGDI PTYVVGVNEE GYTHADTIIS NASCTTNCLA P FVKVLDQK ...String:
KLKVAINGFG RIGRNFLRCW HGRKDSPLDV VVINDTGGVK QASHLLKYDS ILGTFDADVK TAGDSAISVD GKVIKVVSDR NPVNLPWGD MGIDLVIEGT GVFVDRDGAG KHLQAGAKKV LITAPGKGDI PTYVVGVNEE GYTHADTIIS NASCTTNCLA P FVKVLDQK FGIIKGTMTT THSYTGDQRL LDASHRDLRR ARAACLNIVP TSTGAAKAVA LVLPNLKGKL NGIALRVPTP NV SVVDLVV QVSKKTFAEE VNAAFRESAD NELKGILSVC DEPLVSIDFR CTDVSSTIDS SLTMVMGDDM VKVIAWYDNE WGY SQRVVD LADIVANKWQ A

UniProtKB: Glyceraldehyde-3-phosphate dehydrogenase A, chloroplastic

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Macromolecule #3: NICOTINAMIDE-ADENINE-DINUCLEOTIDE

MacromoleculeName: NICOTINAMIDE-ADENINE-DINUCLEOTIDE / type: ligand / ID: 3 / Number of copies: 20 / Formula: NAD
Molecular weightTheoretical: 663.425 Da
Chemical component information

ChemComp-NAD:
NICOTINAMIDE-ADENINE-DINUCLEOTIDE / NAD*YM

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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

BufferpH: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400
VitrificationCryogen name: ETHANE / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 42.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 3.5 µm / Nominal defocus min: 1.5 µm
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 13.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 7532
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

2pkq


Chain - Source name: PDB / Chain - Initial model type: experimental model
Output model

PDB-7q57:
Single Particle Cryo-EM structure of photosynthetic A10B10 glyceraldehyde-3-phospahte dehydrogenase from Spinacia oleracea.

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