[English] 日本語
Yorodumi
- EMDB-45605: Anthoceros agrestis Rubisco assembled with Raf1 Raf2 and BSD2 -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-45605
TitleAnthoceros agrestis Rubisco assembled with Raf1 Raf2 and BSD2
Map dataHornwort Rubisco assembled with Raf1/Raf2/BSD2, main map.
Sample
  • Complex: Rubisco assembled hexadecamer
    • Protein or peptide: Rubisco large subunit
    • Protein or peptide: Rubisco small subunit
  • Ligand: MAGNESIUM ION
  • Ligand: 2-CARBOXYARABINITOL-1,5-DIPHOSPHATE
KeywordsAnthoceros agrestis rubisco / PHOTOSYNTHESIS
Biological speciesAnthoceros agrestis (plant)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsAng WSL / Oh ZG / Li FW / Gunn LH
Funding support United States, 2 items
OrganizationGrant numberCountry
National Science Foundation (NSF, United States)MCB2213840 United States
National Science Foundation (NSF, United States)MCB2213841 United States
CitationJournal: Mol Plant / Year: 2024
Title: Unique biogenesis and kinetics of hornwort Rubiscos revealed by synthetic biology systems.
Authors: Zhen-Guo Oh / Tanner Ashton Robison / Dan-Hong Loh / Warren Shou Leong Ang / Jediael Ng / Fay-Wei Li / Laura Helen Gunn /
Abstract: Hornworts are the only land plants that employ a pyrenoid to optimize Rubisco's CO fixation. Yet, hornwort Rubisco remains poorly characterized. Here we assemble the hornwort Anthoceros agrestis ...Hornworts are the only land plants that employ a pyrenoid to optimize Rubisco's CO fixation. Yet, hornwort Rubisco remains poorly characterized. Here we assemble the hornwort Anthoceros agrestis Rubisco (AaRubisco) using the Arabidopsis thaliana SynBio expression system and observed the formation of stalled intermediates, prompting us to develop a new SynBio system with A. agrestis cognate chaperones. We successfully assembled AaRubisco and Rubisco from three other hornwort species. Unlike A. thaliana Rubisco, AaRubisco assembly is not dependent on RbcX or Raf2. Kinetic characterization reveals that hornwort Rubiscos exhibit a range of catalytic rates (3-10 s), but with similar affinity (∼30 μM) and specificity (∼70) for CO. In other words, hornwort Rubiscos do not comply with the long held canonical catalytic trade-off observed in other land plants, providing experimental support that Rubisco kinetics may be phylogenetically constrained. Unexpectedly, we observed a 50% increase in AaRubisco catalytic rates when RbcX was removed from our SynBio system, without any reduction in specificity. Structural biology, biochemistry and proteomic analysis suggest that subtle differences in Rubisco large subunit interactions, when RbcX is absent during biogenesis, increases the accessibility of active sites and catalytic turnover rate. This study uncovered a previously unknown Rubisco kinetic parameter space and provides a SynBio chassis to expand the survey of other Rubisco kinetics. Our discovery could thus reshape the approaches for engineering Rubisco with superior kinetics.
History
DepositionJul 2, 2024-
Header (metadata) releaseNov 13, 2024-
Map releaseNov 13, 2024-
UpdateNov 20, 2024-
Current statusNov 20, 2024Processing site: RCSB / Status: Released

-
Structure visualization

Supplemental images

emd_45605.png

Downloads & links

-
Map

FileDownload / File: emd_45605.map.gz / Format: CCP4 / Size: 34.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHornwort Rubisco assembled with Raf1/Raf2/BSD2, main map.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.39 Å/pix.
x 208 pix.
= 289.12 Å		1.39 Å/pix.
x 208 pix.
= 289.12 Å		1.39 Å/pix.
x 208 pix.
= 289.12 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.39 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.7
Minimum - Maximum-2.5218267 - 5.0106616
Average (Standard dev.)0.008248672 (±0.22088408)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions208208208
Spacing208208208
CellA=B=C: 289.12 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: Hornwort Rubisco assembled with Raf1/Raf2/BSD2, half map A.

Fileemd_45605_half_map_1.map
AnnotationHornwort Rubisco assembled with Raf1/Raf2/BSD2, half map A.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: Hornwort Rubisco assembled with Raf1/Raf2/BSD2, half map B.

Fileemd_45605_half_map_2.map
AnnotationHornwort Rubisco assembled with Raf1/Raf2/BSD2, half map B.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : Rubisco assembled hexadecamer

EntireName: Rubisco assembled hexadecamer
Components
  • Complex: Rubisco assembled hexadecamer
    • Protein or peptide: Rubisco large subunit
    • Protein or peptide: Rubisco small subunit
  • Ligand: MAGNESIUM ION
  • Ligand: 2-CARBOXYARABINITOL-1,5-DIPHOSPHATE

-
Supramolecule #1: Rubisco assembled hexadecamer

SupramoleculeName: Rubisco assembled hexadecamer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Anthoceros agrestis (plant)
Molecular weightTheoretical: 540 KDa

-
Macromolecule #1: Rubisco large subunit

MacromoleculeName: Rubisco large subunit / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Anthoceros agrestis (plant)
Molecular weightTheoretical: 52.908777 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString: MSPQTETKAG VGFKAGVKDY RLTYYTPDYE TKDTDILAAF RMTPQPGVPP EEAGAAVAAE SSTGTWTTVW TDGLTSLDRY KGRCYDIEP VAGEENQYIA YVAYPLDLFE EGSVTNMFTS IVGNVFGFKA LRALRLEDLR IPPAYSKTFQ GPPHGIQVER D KLNKYGRP ...String:
MSPQTETKAG VGFKAGVKDY RLTYYTPDYE TKDTDILAAF RMTPQPGVPP EEAGAAVAAE SSTGTWTTVW TDGLTSLDRY KGRCYDIEP VAGEENQYIA YVAYPLDLFE EGSVTNMFTS IVGNVFGFKA LRALRLEDLR IPPAYSKTFQ GPPHGIQVER D KLNKYGRP LLGCTIKPKL GLSAKNYGRA VYECLRGGLD FT(KCX)DDENVNS QPFMRWRDRF LFVAEAIFKS QAETGEIK G HYLNATAGTC EEMMKRAQFA RELGMPIVMH DYLTGGFTAN TTLAHYCRDN GLLLHIHRAM HAVIDRQRNH GIHFRVLAK ALRMSGGDHI HSGTVVGKLE GEREVTLGFV DLLRDDYIEK DRSRGIYFTQ DWVSMPGVLP VASGGIHVWH MPALTEIFGD DSVLQFGGG TLGHPWGNAP GAVANRVALE ACVQARNEGR DLAREGNDII REASKWSPEL AAACEVWKEI KFVFETIDTL

-
Macromolecule #2: Rubisco small subunit

MacromoleculeName: Rubisco small subunit / type: protein_or_peptide / ID: 2 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Anthoceros agrestis (plant)
Molecular weightTheoretical: 14.70791 KDa
Recombinant expressionOrganism: Escherichia coli 'BL21-Gold(DE3)pLysS AG' (bacteria)
SequenceString:
MQVWNPIDNP KFETLSYLPP LTDNQIAREI DYMLRNKWIP CLEFDPSGTI TTLPGQPGYY GGRYWTMWKL PMFGCNNAGY VLREIEHCK NAYPGCFIRV LGFDNIRQVQ CCAFIVHKPQ HHHHHH

-
Macromolecule #3: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 3 / Number of copies: 8 / Formula: MG
Molecular weightTheoretical: 24.305 Da

-
Macromolecule #4: 2-CARBOXYARABINITOL-1,5-DIPHOSPHATE

MacromoleculeName: 2-CARBOXYARABINITOL-1,5-DIPHOSPHATE / type: ligand / ID: 4 / Number of copies: 8 / Formula: CAP
Molecular weightTheoretical: 356.115 Da
Chemical component information

ChemComp-CAP:
2-CARBOXYARABINITOL-1,5-DIPHOSPHATE

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration0.5 mg/mL
BufferpH: 8
Component:
ConcentrationFormulaName
20.0 mMC4H11NO3Tris
50.0 mMNaClSodium chloride

Details: 20 mM Tris pH 8.0 50 mM NaCl
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

-
Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Average exposure time: 4.8 sec. / Average electron dose: 65.0 e/Å2 / Details: Images were collected over 50 movie frames
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 63000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

+
Image processing

Particle selectionNumber selected: 3252384
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

1gk8


Details: Rigid body docking using 1GK8 as the starting model
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: D4 (2x4 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4) / Software - details: Homogeneous refinement / Number images used: 222666
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4)
Final 3D classificationNumber classes: 4 / Avg.num./class: 183580 / Software - Name: cryoSPARC (ver. 4) / Software - details: 3D classification tool

-
Atomic model buiding 1

Initial modelPDB ID:

1gk8


Chain - Chain ID: all / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-9chz:
Anthoceros agrestis Rubisco assembled with Raf1 Raf2 and BSD2

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more