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- EMDB-61158: Cryo-EM structure of the rice isoamylase ISA1 dimer -

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Basic information

Entry
Database: EMDB / ID: EMD-61158
TitleCryo-EM structure of the rice isoamylase ISA1 dimer
Map data
Sample
  • Complex: ISA1 dimer
    • Protein or peptide: Isoamylase 1, chloroplastic
Keywordsisoamylase ISA1-ISA2 / HYDROLASE
Function / homology
Function and homology information


chloroplast isoamylase complex / isoamylase / isoamylase complex / isoamylase activity / amylopectin biosynthetic process / starch biosynthetic process / starch catabolic process
Similarity search - Function
: / Isoamylase 1-3, C-terminal / Glycogen debranching enzyme GlgX/isoamylase, N-terminal Early set domain / Glycoside hydrolase, family 13, N-terminal / Carbohydrate-binding module 48 (Isoamylase N-terminal domain) / Alpha amylase, catalytic domain / Glycosyl hydrolase, family 13, catalytic domain / Alpha-amylase domain / Glycosyl hydrolase, all-beta / Immunoglobulin E-set ...: / Isoamylase 1-3, C-terminal / Glycogen debranching enzyme GlgX/isoamylase, N-terminal Early set domain / Glycoside hydrolase, family 13, N-terminal / Carbohydrate-binding module 48 (Isoamylase N-terminal domain) / Alpha amylase, catalytic domain / Glycosyl hydrolase, family 13, catalytic domain / Alpha-amylase domain / Glycosyl hydrolase, all-beta / Immunoglobulin E-set / Glycoside hydrolase superfamily / Immunoglobulin-like fold
Similarity search - Domain/homology
Isoamylase 1, chloroplastic
Similarity search - Component
Biological speciesOryza sativa Japonica Group (Japanese rice)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.7 Å
AuthorsGuan ZY / Yan JJ
Funding support China, 1 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Nat Commun / Year: 2025
Title: Amylopectin branch trimming and biosynthesis elucidated by the rice isoamylase ISA1-ISA2 heterocomplex.
Authors: Rong Fan / Zeyuan Guan / Guanghong Zhou / Xi Yang / Fei Zhang / Menglong Wu / Xuecui Wang / Jian Liu / Pei Chen / Yanjun Liu / Delin Zhang / Ping Yin / Junjie Yan /
Abstract: Amylopectin, the primary form of starch in plant leaves, seeds and tubers, features a tree-like architecture with branched glucose chains. Excess branches result in the formation of soluble ...Amylopectin, the primary form of starch in plant leaves, seeds and tubers, features a tree-like architecture with branched glucose chains. Excess branches result in the formation of soluble phytoglycogen instead of starch granules. In higher plants and green algae, the debranching enzyme isoamylase ISA1 forms either homomultimer or hetero-multimer with ISA2 to facilitate branch trimming and starch granule formation, but the molecular basis remains largely unknown. In this study, we reconstitute the rice OsISA1-ISA2 complex in vitro and determine the cryo-EM structures of the OsISA1 homodimer, as well as the malto-oligosaccharide (MOS)-free and MOS-bound OsISA1-ISA2 heterocomplex. The OsISA1 dimer shows a tail-to-tail rod-like architecture, whereas the OsISA1-ISA2 complex mainly exhibits as a trimer, with OsISA2 flanking on the N-terminal segments of the dimeric OsISA1. Combined with comprehensive biochemical analyses, these structural data elucidate the organization of the ISA1-ISA2 heterocomplex in higher plants and demonstrate how ISA1 and ISA2 cooperate during amylopectin biosynthesis.
History
DepositionAug 14, 2024-
Header (metadata) releaseMay 28, 2025-
Map releaseMay 28, 2025-
UpdateJul 16, 2025-
Current statusJul 16, 2025Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_61158.png

Downloads & links

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Map

FileDownload / File: emd_61158.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 400 pix.
= 336. Å		0.84 Å/pix.
x 400 pix.
= 336. Å		0.84 Å/pix.
x 400 pix.
= 336. 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: 0.35
Minimum - Maximum-2.0481148 - 2.8420498
Average (Standard dev.)-0.0003363276 (±0.070477545)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 336.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_61158_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_61158_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 : ISA1 dimer

EntireName: ISA1 dimer
Components
  • Complex: ISA1 dimer
    • Protein or peptide: Isoamylase 1, chloroplastic

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Supramolecule #1: ISA1 dimer

SupramoleculeName: ISA1 dimer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Oryza sativa Japonica Group (Japanese rice)

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Macromolecule #1: Isoamylase 1, chloroplastic

MacromoleculeName: Isoamylase 1, chloroplastic / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: isoamylase
Source (natural)Organism: Oryza sativa Japonica Group (Japanese rice)
Molecular weightTheoretical: 87.281594 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MGSSHHHHHH SSGLVPRGSH SDEVDAHMSV ASAVEVGVGE DEEEGVEEEE EEVEAVVMPE RYALGGACRV LAGMPAPLGA TALDGGVNF AVYSAGASAA SLCLFTPDDL EADEVTEEVP LDPLFNRTGN VWHVFIEGEL HNMLYGYRFD GMFAPHCGQY F DVSNVVVD ...String:
MGSSHHHHHH SSGLVPRGSH SDEVDAHMSV ASAVEVGVGE DEEEGVEEEE EEVEAVVMPE RYALGGACRV LAGMPAPLGA TALDGGVNF AVYSAGASAA SLCLFTPDDL EADEVTEEVP LDPLFNRTGN VWHVFIEGEL HNMLYGYRFD GMFAPHCGQY F DVSNVVVD PYAKAVISRG EYGVPGPGGD CWPQMAGMIP LPYSTFDWQG DLPLRYPQKD LVIYEMHLRG FTKHSSSNVE HP GTYIGAI SKLDYLKELG VNCVELMPCH EFNELEYFSC SSKMNFWGYS TINFFSPMIR YSSGGIRNCG RDAINEFKTF VRE AHKRGI EVIMDVVFNH TAEGNEKGPI LSFRGIDNST YYMLAPKGEF YNYSGCGNTF NCNHPVVREF IVDCLRYWVT EMHV DGFRF DLASIMTRGC SLWDPVNVYG SPVEGDMTTT GTPLATPPLI DMISNDPILG DVKLIAEAWD AGGLYQVGQF PHWKI WSEW NGKYRDIVRQ FIKGTDGFAG GFAECLCGSP HLYQAGGRKP WHSINFVCAH DGFTLADLVT YNKKYNSSNG EDNRDG ENH NLSWNCGEEG EFAGLSVKRL RKRQMRNFFV SLMVSQGVPM FYMGDEYGHT KGGNNNTYCH DHYVNYFRWD KKEESSD LQ RFCSLMTKFR KQCESLGLAD FPTAQRLHWH GHQPGKPDWS ETSRFVAFST KDETKGEIYV AFNASHLPAV VGLPERPG Y RWEPLVDTGK PAPYDFLTDD LPDRAHAVHL FSHFLNSNLY PMLSYSSIIL ELQPDD

UniProtKB: Isoamylase 1, chloroplastic

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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

Concentration0.75 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / 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 / Nominal defocus max: 1.6 µm / Nominal defocus min: 1.2 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: NONE
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 104741
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: PROJECTION MATCHING

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