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- EMDB-10235: Cryo-electron microscopy structure of a RbcL-Raf1 supercomplex fr... -

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Basic information

Entry
Database: EMDB / ID: EMD-10235
TitleCryo-electron microscopy structure of a RbcL-Raf1 supercomplex from Synechococcus elongatus PCC 7942
Map dataCryo-electron microscopy structure of a RbcL8-Raf18 supercomplex from the cyanobacterium Synechococcus elongatus 7942
Sample
  • Complex: RbcL-Raf1 supercomplex of Synechococcus elongatus 7942
    • Protein or peptide: Ribulose bisphosphate carboxylase large chain
    • Protein or peptide: Rubisco accumulation factor 1 (RAF1)
Function / homology
Function and homology information


ribulose bisphosphate carboxylase complex assembly / photorespiration / carboxysome / ribulose-bisphosphate carboxylase / carbon fixation / ribulose-bisphosphate carboxylase activity / reductive pentose-phosphate cycle / photosynthesis / monooxygenase activity / magnesium ion binding / cytoplasm
Similarity search - Function
Rubisco accumulation factor 1 / Rubisco accumulation factor 1, helix turn helix domain / Rubisco accumulation factor 1, C-terminal / Rubisco accumulation factor 1, alpha helical domain / Rubisco Assembly chaperone C-terminal domain / Rubisco accumulation factor 1 alpha helical domain / Rubisco accumulation factor 1 helix turn helix domain / Ribulose bisphosphate carboxylase large subunit, type I / Ribulose bisphosphate carboxylase, large chain, active site / Ribulose bisphosphate carboxylase large chain active site. ...Rubisco accumulation factor 1 / Rubisco accumulation factor 1, helix turn helix domain / Rubisco accumulation factor 1, C-terminal / Rubisco accumulation factor 1, alpha helical domain / Rubisco Assembly chaperone C-terminal domain / Rubisco accumulation factor 1 alpha helical domain / Rubisco accumulation factor 1 helix turn helix domain / Ribulose bisphosphate carboxylase large subunit, type I / Ribulose bisphosphate carboxylase, large chain, active site / Ribulose bisphosphate carboxylase large chain active site. / Ribulose bisphosphate carboxylase, large subunit, ferrodoxin-like N-terminal / Ribulose bisphosphate carboxylase large chain, N-terminal domain / Ribulose bisphosphate carboxylase, large subunit, C-terminal / RuBisCO / Ribulose bisphosphate carboxylase, large subunit, C-terminal domain superfamily / RuBisCO large subunit, N-terminal domain superfamily / Ribulose bisphosphate carboxylase large chain, catalytic domain
Similarity search - Domain/homology
Ribulose bisphosphate carboxylase large chain / RuBisCO accumulation factor 1
Similarity search - Component
Biological speciesSynechococcus elongatus PCC 7942 (bacteria) / Synechococcus elongatus PCC 7942 = FACHB-805 (bacteria) / Synechococcus elongatus (strain PCC 7942 / FACHB-805) (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsHuang F / Kong W-W / Sun Y / Chen T / Dykes GF / Jiang YL / Liu LN
Funding support United Kingdom, 4 items
OrganizationGrant numberCountry
Royal SocietyURF/R/180030 United Kingdom
Biotechnology and Biological Sciences Research Council (BBSRC)BB/M024202/1 United Kingdom
Royal SocietyUF120411 United Kingdom
Leverhulme TrustECF-2016-778 United Kingdom
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: Rubisco accumulation factor 1 (Raf1) plays essential roles in mediating Rubisco assembly and carboxysome biogenesis.
Authors: Fang Huang / Wen-Wen Kong / Yaqi Sun / Taiyu Chen / Gregory F Dykes / Yong-Liang Jiang / Lu-Ning Liu /
Abstract: Carboxysomes are membrane-free organelles for carbon assimilation in cyanobacteria. The carboxysome consists of a proteinaceous shell that structurally resembles virus capsids and internal enzymes ...Carboxysomes are membrane-free organelles for carbon assimilation in cyanobacteria. The carboxysome consists of a proteinaceous shell that structurally resembles virus capsids and internal enzymes including ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), the primary carbon-fixing enzyme in photosynthesis. The formation of carboxysomes requires hierarchical self-assembly of thousands of protein subunits, initiated from Rubisco assembly and packaging to shell encapsulation. Here we study the role of Rubisco assembly factor 1 (Raf1) in Rubisco assembly and carboxysome formation in a model cyanobacterium, PCC7942 (Syn7942). Cryo-electron microscopy reveals that Raf1 facilitates Rubisco assembly by mediating RbcL dimer formation and dimer-dimer interactions. Syn7942 cells lacking Raf1 are unable to form canonical intact carboxysomes but generate a large number of intermediate assemblies comprising Rubisco, CcaA, CcmM, and CcmN without shell encapsulation and a low abundance of carboxysome-like structures with reduced dimensions and irregular shell shapes and internal organization. As a consequence, the Raf1-depleted cells exhibit reduced Rubisco content, CO-fixing activity, and cell growth. Our results provide mechanistic insight into the chaperone-assisted Rubisco assembly and biogenesis of carboxysomes. Advanced understanding of the biogenesis and stepwise formation process of the biogeochemically important organelle may inform strategies for heterologous engineering of functional CO-fixing modules to improve photosynthesis.
History
DepositionAug 18, 2019-
Header (metadata) releaseAug 5, 2020-
Map releaseAug 5, 2020-
UpdateAug 5, 2020-
Current statusAug 5, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6smh
  • Surface level: 0.015
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10235.png

Downloads & links

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Map

FileDownload / File: emd_10235.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryo-electron microscopy structure of a RbcL8-Raf18 supercomplex from the cyanobacterium Synechococcus elongatus 7942
Voxel sizeX=Y=Z: 1.25 Å
Density
Contour LevelBy AUTHOR: 0.015 / Movie #1: 0.015
Minimum - Maximum-0.04692439 - 0.07288982
Average (Standard dev.)0.00019264383 (±0.004175179)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 300.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.251.251.25
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z300.000300.000300.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.0470.0730.000

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Supplemental data

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Sample components

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Entire : RbcL-Raf1 supercomplex of Synechococcus elongatus 7942

EntireName: RbcL-Raf1 supercomplex of Synechococcus elongatus 7942
Components
  • Complex: RbcL-Raf1 supercomplex of Synechococcus elongatus 7942
    • Protein or peptide: Ribulose bisphosphate carboxylase large chain
    • Protein or peptide: Rubisco accumulation factor 1 (RAF1)

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Supramolecule #1: RbcL-Raf1 supercomplex of Synechococcus elongatus 7942

SupramoleculeName: RbcL-Raf1 supercomplex of Synechococcus elongatus 7942
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Details: cryoEM structure of a RbcL8-Raf18 complex from Synechococcus elongatus 7942
Source (natural)Organism: Synechococcus elongatus PCC 7942 (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Macromolecule #1: Ribulose bisphosphate carboxylase large chain

MacromoleculeName: Ribulose bisphosphate carboxylase large chain / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO / EC number: ribulose-bisphosphate carboxylase
Source (natural)Organism: Synechococcus elongatus PCC 7942 = FACHB-805 (bacteria)
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: LTYYTPDYTP KDTDLLAAFR FSPQPGVPAP EAGAAIAAES STGTWTTVWT DLLTDMDRYD GKCYHIEPVQ GEENSYFAFI ADPLDLFEE GSVTNILTSI VGNVFGFKAI RSLRLEDIRF PVALVKTFQG PPHGIQVERD LLNKYGRPML GCTIKPKLGL S AKNYGRAV ...String:
LTYYTPDYTP KDTDLLAAFR FSPQPGVPAP EAGAAIAAES STGTWTTVWT DLLTDMDRYD GKCYHIEPVQ GEENSYFAFI ADPLDLFEE GSVTNILTSI VGNVFGFKAI RSLRLEDIRF PVALVKTFQG PPHGIQVERD LLNKYGRPML GCTIKPKLGL S AKNYGRAV YECLRGGLDF TKDDENINSQ PFQRWRDRFL FVADAIHKSQ AETGEIKGHY LNVTAPTCEE MMKRAEFAKE LG MPIIMHD FLTAGFTANT TLAKWCRDNG VLLHIHRAMH AVIDRQRNHG IHFRVLAKCL RLSGGDHLHS GTVVGKLEGD KAS TLGFVD LMREDHIEAD RSRGVFFTQD WASMPGVLPV ASGGIHVWHM PALVEIFGDD SVLQFGGGTL GHPWGNAPGA TANR VALEA CVQARNEGRD LYREGGDILR EAGKWSPELA AALDLWKEIK FE

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Macromolecule #2: Rubisco accumulation factor 1 (RAF1)

MacromoleculeName: Rubisco accumulation factor 1 (RAF1) / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Synechococcus elongatus (strain PCC 7942 / FACHB-805) (bacteria)
SequenceString:
ERQELLGQLR RKEGRWLAWA RACQTLLKNG LNPQTLFEAT GFEPIQQNQI TVAMQVYDSI LRQDPPAHVR ETYQEWGSDL LYELRELDQ EQRSLCAQLA LERKLDADQI REVAKATKDF CRLPKQPENF DRHPGDAVAH QCWRLAQERT DLTERSRLIA R GLQFAQSA GARALIEALL LDLSGVPSRK

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation state3D array

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Sample preparation

BufferpH: 8
VitrificationCryogen name: NITROGEN

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Electron microscopy

MicroscopeFEI TECNAI 20
Electron beamAcceleration voltage: 200 kV / Electron source: OTHER
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 25.0 e/Å2

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6kkn

Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.3 Å / Resolution method: OTHER / Number images used: 16022
FSC plot (resolution estimation)

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