+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-33887 | |||||||||
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Title | Rubisco from Phaeodactylum tricornutum bound to PYCO1(452-592) | |||||||||
Map data | Composite map of multibody refinement. Composite from EMD-35158 and EMD-35159. Consensus maps are deposited at: D_1300034757; EMD-35166 | |||||||||
Sample |
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Keywords | Rubisco / phase separation / rubisco linker protein / condensation / pyrenoid / phaeodactylum tricornutum / PHOTOSYNTHESIS | |||||||||
Function / homology | Function and homology information ribulose-bisphosphate carboxylase / ribulose-bisphosphate carboxylase activity / reductive pentose-phosphate cycle / monooxygenase activity / chloroplast / magnesium ion binding Similarity search - Function | |||||||||
Biological species | Phaeodactylum tricornutum (Diatom) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.0 Å | |||||||||
Authors | Oh ZG / Ang WSL / Bhushan S / Mueller-Cajar O | |||||||||
Funding support | Singapore, 2 items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: A linker protein from a red-type pyrenoid phase separates with Rubisco via oligomerizing sticker motifs. Authors: Zhen Guo Oh / Warren Shou Leong Ang / Cheng Wei Poh / Soak-Kuan Lai / Siu Kwan Sze / Hoi-Yeung Li / Shashi Bhushan / Tobias Wunder / Oliver Mueller-Cajar / Abstract: The slow kinetics and poor substrate specificity of the key photosynthetic CO-fixing enzyme Rubisco have prompted the repeated evolution of Rubisco-containing biomolecular condensates known as ...The slow kinetics and poor substrate specificity of the key photosynthetic CO-fixing enzyme Rubisco have prompted the repeated evolution of Rubisco-containing biomolecular condensates known as pyrenoids in the majority of eukaryotic microalgae. Diatoms dominate marine photosynthesis, but the interactions underlying their pyrenoids are unknown. Here, we identify and characterize the Rubisco linker protein PYCO1 from . PYCO1 is a tandem repeat protein containing prion-like domains that localizes to the pyrenoid. It undergoes homotypic liquid-liquid phase separation (LLPS) to form condensates that specifically partition diatom Rubisco. Saturation of PYCO1 condensates with Rubisco greatly reduces the mobility of droplet components. Cryo-electron microscopy and mutagenesis data revealed the sticker motifs required for homotypic and heterotypic phase separation. Our data indicate that the PYCO1-Rubisco network is cross-linked by PYCO1 stickers that oligomerize to bind to the small subunits lining the central solvent channel of the Rubisco holoenzyme. A second sticker motif binds to the large subunit. Pyrenoidal Rubisco condensates are highly diverse and tractable models of functional LLPS. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_33887.map.gz | 4.6 MB | EMDB map data format | |
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Header (meta data) | emd-33887-v30.xml emd-33887.xml | 24.8 KB 24.8 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_33887_fsc.xml | 9.2 KB | Display | FSC data file |
Images | emd_33887.png | 196.7 KB | ||
Others | emd_33887_additional_1.map.gz emd_33887_additional_2.map.gz | 84.6 KB 4.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-33887 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-33887 | HTTPS FTP |
-Validation report
Summary document | emd_33887_validation.pdf.gz | 516.8 KB | Display | EMDB validaton report |
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Full document | emd_33887_full_validation.pdf.gz | 516.3 KB | Display | |
Data in XML | emd_33887_validation.xml.gz | 10.9 KB | Display | |
Data in CIF | emd_33887_validation.cif.gz | 14.3 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-33887 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-33887 | HTTPS FTP |
-Related structure data
Related structure data | 7yk5MC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_33887.map.gz / Format: CCP4 / Size: 67 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Composite map of multibody refinement. Composite from EMD-35158 and EMD-35159. Consensus maps are deposited at: D_1300034757; EMD-35166 | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.858 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Additional map: Map of peptide attached to Rubisco. Second part...
File | emd_33887_additional_1.map | ||||||||||||
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Annotation | Map of peptide attached to Rubisco. Second part of composite map. Also deposited as D_1300034756; EMD-35159 with the half maps | ||||||||||||
Projections & Slices |
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Density Histograms |
-Additional map: Map of Rubisco (main protein). First part of...
File | emd_33887_additional_2.map | ||||||||||||
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Annotation | Map of Rubisco (main protein). First part of composite map. Also deposited as D_1300034754; EMD-35158 with the half maps. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : Rubisco from Phaeodactylum tricornutum bound to linker protein PYCO1
Entire | Name: Rubisco from Phaeodactylum tricornutum bound to linker protein PYCO1 |
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Components |
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-Supramolecule #1: Rubisco from Phaeodactylum tricornutum bound to linker protein PYCO1
Supramolecule | Name: Rubisco from Phaeodactylum tricornutum bound to linker protein PYCO1 type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4 Details: Rubisco purified from source organism, PYCO1 purified from E. coli. |
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Source (natural) | Organism: Phaeodactylum tricornutum (Diatom) |
Molecular weight | Theoretical: 550 kDa/nm |
-Macromolecule #1: Ribulose bisphosphate carboxylase large chain
Macromolecule | Name: Ribulose bisphosphate carboxylase large chain / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO / EC number: ribulose-bisphosphate carboxylase |
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Source (natural) | Organism: Phaeodactylum tricornutum (Diatom) |
Molecular weight | Theoretical: 54.244527 KDa |
Sequence | String: MSQSVSERTR IKSDRYESGV IPYAKMGYWD AAYAVKNTDV LALFRIT(HYP)QP GVDPVEAAAA VAGESSTATW TVVWTD LLT ACDRYRAKAY RVDPVPNTTD QYFAFIAYE(CSO) DLFEEGSLAN LTASIIGNVF GFKAVSALRL EDMRIPHSYL (LYO)TFQG(HYP)ATG ...String: MSQSVSERTR IKSDRYESGV IPYAKMGYWD AAYAVKNTDV LALFRIT(HYP)QP GVDPVEAAAA VAGESSTATW TVVWTD LLT ACDRYRAKAY RVDPVPNTTD QYFAFIAYE(CSO) DLFEEGSLAN LTASIIGNVF GFKAVSALRL EDMRIPHSYL (LYO)TFQG(HYP)ATG VIVERERLNK YGIPL(HLU)GATV KPKLGLSGKN YGRVVYEGL(LYO) GGLDFL(KCX)DDE N INSQPFMR WRERFLYCME GINRASAATG ETKGSYLNIT AGTMEEVYKR AEYAKTVGSI VVMIDLVMGY TAIQSAAIWA RD NDLILHL HRAGNSTYAR QKNHGINFRV ICKWMRMCGV DHIHAGTVVG KLEGDPLMI(M3L) GFYDTLLLTH LNVNLPYGI FFEMTWASLR KCMPVASGGI HCGQMHQLVH YLGDDVVLQF GGGTIGHPDG IQAGATANRV ALEAMILARN EGADYFNSDI GPQILRNAA KTCGPLQTAL DLWKDISFNY TSTDTSDFSV TPTANV |
-Macromolecule #2: Multifunctional fusion protein
Macromolecule | Name: Multifunctional fusion protein / type: protein_or_peptide / ID: 2 / Number of copies: 8 / Enantiomer: LEVO |
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Source (natural) | Organism: Phaeodactylum tricornutum (Diatom) |
Molecular weight | Theoretical: 16.039053 KDa |
Sequence | String: MRLTQGCFSF LPDLTDQQIE KQIAYCITKG WAMNVEWTDD PHPRNSYWEL WGLPLFDVKD PASVMFELRE ARKSCAAGYI RINAFNAAY GTESCVMSFI VNRPSNEPGF YLERQELEGR RIAYTTKSYS VQANPEGGRY |
-Macromolecule #3: PYCO1 SSU binding motif
Macromolecule | Name: PYCO1 SSU binding motif / type: protein_or_peptide / ID: 3 / Number of copies: 4 / Enantiomer: LEVO |
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Source (natural) | Organism: Phaeodactylum tricornutum (Diatom) |
Molecular weight | Theoretical: 875.973 Da |
Recombinant expression | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Sequence | String: KWSPRGGS |
-Macromolecule #4: PYCO1 LSU binding motif
Macromolecule | Name: PYCO1 LSU binding motif / type: protein_or_peptide / ID: 4 / Number of copies: 8 / Enantiomer: LEVO |
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Source (natural) | Organism: Phaeodactylum tricornutum (Diatom) |
Molecular weight | Theoretical: 993.051 Da |
Recombinant expression | Organism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria) |
Sequence | String: AAEWGSMNQ |
-Macromolecule #5: 2-CARBOXYARABINITOL-1,5-DIPHOSPHATE
Macromolecule | Name: 2-CARBOXYARABINITOL-1,5-DIPHOSPHATE / type: ligand / ID: 5 / Number of copies: 8 / Formula: CAP |
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Molecular weight | Theoretical: 356.115 Da |
Chemical component information | ChemComp-CAP: |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.5 mg/mL | |||||||||
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Buffer | pH: 8 Component:
Details: 20 mM Tris pH 8.0 20 mM NaCl | |||||||||
Grid | Model: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 20 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.6 kPa | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: Blotted for 2 sec with blot force of 1.. | |||||||||
Details | 0.5 mg/mL Rubisco incubated with 21.9 uM of PYCO1(452-592). |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV / Details: Gatan EF |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 8861 / Average exposure time: 5.0 sec. / Average electron dose: 65.0 e/Å2 Details: Images were collected in movie mode at 10 frames per second. |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 165000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |