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- EMDB-56441: CryoEM map of chloroplastic photosynthetic NADP(+)-dependent mali... -

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Basic information

Entry
Database: EMDB / ID: EMD-56441
TitleCryoEM map of chloroplastic photosynthetic NADP(+)-dependent malic enzyme mutant (G200R) at pH 8
Map data
Sample
  • Complex: Mutant C4-NADP-malic enzyme (G200R)
    • Protein or peptide: C4-NADP malic enzyme
Keywordsdecarboxylation / malate / C4 pathway / C4 carbon concentrating mechanism / PHOTOSYNTHESIS
Biological speciesZea mays (maize)
Methodsingle particle reconstruction / cryo EM / Resolution: 5.5 Å
AuthorsDrakonaki A / Gatsogiannis C
Funding support Germany, 4 items
OrganizationGrant numberCountry
German Research Foundation (DFG)441941117 Germany
German Research Foundation (DFG)424228829 Germany
Other governmentPICT-2019-00079
German Research Foundation (DFG)496113311 Germany
CitationJournal: Mol Biol Evol / Year: 2026
Title: A milestone in C4 carbon concentration mechanism evolution: structural remodeling of NADP-malic enzyme in Poaceae.
Authors: Jonas M Böhm / Simone Willms / Oja Ferrao / Martin Buitrago-Arango / Meike Hüdig / Gereon Poschmann / Nazanin Fazelnia / Luitgard Nagel-Steger / Sebastián Klinke / Athina Drakonaki / ...Authors: Jonas M Böhm / Simone Willms / Oja Ferrao / Martin Buitrago-Arango / Meike Hüdig / Gereon Poschmann / Nazanin Fazelnia / Luitgard Nagel-Steger / Sebastián Klinke / Athina Drakonaki / Christos Gatsogiannis / Marcos A Tronconi / Clarisa E Alvarez / Veronica G Maurino /
Abstract: The evolution of C4 photosynthesis required extensive modification of ancestral enzymes enabling the development of an efficient carbon concentrating mechanism. A key example is NADP-malic enzyme ...The evolution of C4 photosynthesis required extensive modification of ancestral enzymes enabling the development of an efficient carbon concentrating mechanism. A key example is NADP-malic enzyme (NADP-ME), which, in maize and sorghum-members of the same C4 lineage-underwent gene duplication and neofunctionalization, resulting in 2 plastidic isoforms with distinct oligomeric states: a tetrameric C4-specific isoform and a dimeric housekeeping (nonC4) isoform. In this study, we resolve the structural basis of this oligomeric divergence using X-ray crystallography, cryo-electron microscopy, and molecular modeling combined with targeted biochemical analysis. Our findings demonstrate that the N-terminal region of nonC4-NADP-ME is involved in its oligomeric organization, whereas a suite of adaptive substitutions at the dimer interface drives the transition to the stable tetramer characteristic of the C4 isoform. Moreover, the C-terminal region stabilizes the oligomeric states of C4- and nonC4-NADP-ME through specific interactions with adaptive residues. We propose that tetramerization mitigates aggregation at the high expression levels demanded by the C4 cycle and likely creates a scaffold for the emergence of regulatory properties. Collectively, the data show that remodeling of terminal domains and inter-subunit interfaces rewires the quaternary architecture of the enzymes, illustrating how subtle structural changes can drive the evolution of complex innovations such as C4 photosynthesis.
History
DepositionJan 21, 2026-
Header (metadata) releaseMay 6, 2026-
Map releaseMay 6, 2026-
UpdateMay 6, 2026-
Current statusMay 6, 2026Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_56441.png

Downloads & links

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Map

FileDownload / File: emd_56441.map.gz / Format: CCP4 / Size: 42.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.14 Å/pix.
x 224 pix.
= 255.808 Å		1.14 Å/pix.
x 224 pix.
= 255.808 Å		1.14 Å/pix.
x 224 pix.
= 255.808 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.142 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0968
Minimum - Maximum-0.17463511 - 0.30505705
Average (Standard dev.)0.0005980595 (±0.010422073)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions224224224
Spacing224224224
CellA=B=C: 255.80798 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_56441_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_56441_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 : Mutant C4-NADP-malic enzyme (G200R)

EntireName: Mutant C4-NADP-malic enzyme (G200R)
Components
  • Complex: Mutant C4-NADP-malic enzyme (G200R)
    • Protein or peptide: C4-NADP malic enzyme

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Supramolecule #1: Mutant C4-NADP-malic enzyme (G200R)

SupramoleculeName: Mutant C4-NADP-malic enzyme (G200R) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Zea mays (maize)

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Macromolecule #1: C4-NADP malic enzyme

MacromoleculeName: C4-NADP malic enzyme / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Zea mays (maize)
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MGHHHHHHHH HHSSGHIEGR HAMVSNAET ETEKEQEEAA AASEELPVMP WATSVASGYT LLRDPHHNKG LAFTEEERDG HYLRGLLPPA VLSQELQIKK F MNTLRQYQ TPLQRYIAMM NLQETDERLF YKLLIDNVVE LLPFVYTPTV GEACQKYGSI FRRPQGLYVS ...String:
MGHHHHHHHH HHSSGHIEGR HAMVSNAET ETEKEQEEAA AASEELPVMP WATSVASGYT LLRDPHHNKG LAFTEEERDG HYLRGLLPPA VLSQELQIKK F MNTLRQYQ TPLQRYIAMM NLQETDERLF YKLLIDNVVE LLPFVYTPTV GEACQKYGSI FRRPQGLYVS LKDKGKVLEV LRNWPHRNIQ VICVTDGERI LG LGDLGCQ GMGIPVGKLA LYTALGGVDP SVCLPITIDV GTNNEKLLND EFYIGLRQKR ATGEEYDELI EEFMSAVKQF YGEKVLIQFE DFANHNAFDL LEK YSKSHL VFNDDIQGTA SVVLAGLLAA LKMVGGTLAE QTYLFLGAGE AGTGIAELIA LEISKQTNAP IEECRKKVWL VDSKGLIVDS RKGSLQPFKK PWAH EHEPL KTLYDAVQSI KPTVLIGTSG VGRTFTKEII EAMSSFNERP IIFSLSNPTS HSECTAEQAY TWSQGRSIFA SGSPFAPVEY EGKTFVPGQS NNAYI FPGL GLGLVISGAV RVHEDMLLAA SKALADQATQ DNFEKGSIFP PFTSIRKISA HIAAAVAAKA YELGLATRLP PPSDLVKYAE NCMYTPVYRN YR

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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

Concentration0.5 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
5.0 mMMgCl2Magnesium chloride
40.0 mMPyruvate
2.0 mMNADPNicotinamide Adenine Dinucleotide Phosphate

Details: 20 mM Tris-HCl
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 90 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 286.15 K / Instrument: FEI VITROBOT MARK IV
DetailsThis sample was monodisperse.

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 10 eV
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.6 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 215000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 393438
Details: These particles were picked using Blob picker in cryoSPARC.
CTF correctionSoftware - Name: cryoSPARC (ver. v4.6.2) / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 5.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v4.6.2) / Number images used: 168186
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.6.2)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. v4.6.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: RIGID BODY FIT

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