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- EMDB-9192: Cryo-electron microscopy structure of Plasmodium falciparum Rh5/C... -

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Basic information

Entry
Database: EMDB / ID: EMD-9192
TitleCryo-electron microscopy structure of Plasmodium falciparum Rh5/CyRPA/Ripr invasion complex
Map data
Sample
  • Complex: PfRh5/CyRPA/PfRipr complex
    • Protein or peptide: Cysteine-rich protective antigen
    • Protein or peptide: Reticulocyte binding protein 5
    • Protein or peptide: PfRipr
    • Protein or peptide: PfRipr
Function / homology
Function and homology information


microneme lumen / microneme / symbiont entry into host / apical part of cell / cytoplasmic vesicle / host extracellular space / host cell plasma membrane / protein-containing complex / extracellular region / plasma membrane
Similarity search - Function
Rh5 coiled-coil domain / Rh5 coiled-coil domain / Cysteine-rich protective antigen 6 bladed domain / Cysteine-Rich Protective Antigen 6 bladed domain
Similarity search - Domain/homology
Reticulocyte binding protein 5 / Cysteine-rich protective antigen
Similarity search - Component
Biological speciesPlasmodium falciparum 3D7 (eukaryote) / Plasmodium falciparum (isolate 3D7) (eukaryote) / Plasmodium falciparum (malaria parasite P. falciparum)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.17 Å
AuthorsWilson W / Zhiheng Y / Cowman AF
Funding support Australia, 1 items
OrganizationGrant numberCountry
National Health and Medical Research Council (NHMRC, Australia)GNT637406 Australia
CitationJournal: Nature / Year: 2019
Title: Structure of Plasmodium falciparum Rh5-CyRPA-Ripr invasion complex.
Authors: Wilson Wong / Rick Huang / Sebastien Menant / Chuan Hong / Jarrod J Sandow / Richard W Birkinshaw / Julie Healer / Anthony N Hodder / Usheer Kanjee / Christopher J Tonkin / Denise Heckmann / ...Authors: Wilson Wong / Rick Huang / Sebastien Menant / Chuan Hong / Jarrod J Sandow / Richard W Birkinshaw / Julie Healer / Anthony N Hodder / Usheer Kanjee / Christopher J Tonkin / Denise Heckmann / Vladislav Soroka / Teit Max Moscote Søgaard / Thomas Jørgensen / Manoj T Duraisingh / Peter E Czabotar / Willem A de Jongh / Wai-Hong Tham / Andrew I Webb / Zhiheng Yu / Alan F Cowman /
Abstract: Plasmodium falciparum causes the severe form of malaria that has high levels of mortality in humans. Blood-stage merozoites of P. falciparum invade erythrocytes, and this requires interactions ...Plasmodium falciparum causes the severe form of malaria that has high levels of mortality in humans. Blood-stage merozoites of P. falciparum invade erythrocytes, and this requires interactions between multiple ligands from the parasite and receptors in hosts. These interactions include the binding of the Rh5-CyRPA-Ripr complex with the erythrocyte receptor basigin, which is an essential step for entry into human erythrocytes. Here we show that the Rh5-CyRPA-Ripr complex binds the erythrocyte cell line JK-1 significantly better than does Rh5 alone, and that this binding occurs through the insertion of Rh5 and Ripr into host membranes as a complex with high molecular weight. We report a cryo-electron microscopy structure of the Rh5-CyRPA-Ripr complex at subnanometre resolution, which reveals the organization of this essential invasion complex and the mode of interactions between members of the complex, and shows that CyRPA is a critical mediator of complex assembly. Our structure identifies blades 4-6 of the β-propeller of CyRPA as contact sites for Rh5 and Ripr. The limited contacts between Rh5-CyRPA and CyRPA-Ripr are consistent with the dissociation of Rh5 and Ripr from CyRPA for membrane insertion. A comparision of the crystal structure of Rh5-basigin with the cryo-electron microscopy structure of Rh5-CyRPA-Ripr suggests that Rh5 and Ripr are positioned parallel to the erythrocyte membrane before membrane insertion. This provides information on the function of this complex, and thereby provides insights into invasion by P. falciparum.
History
DepositionOct 8, 2018-
Header (metadata) releaseOct 24, 2018-
Map releaseDec 12, 2018-
UpdateDec 18, 2019-
Current statusDec 18, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.034
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.034
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6mpv
  • Surface level: 0.034
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6mpv
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_9192.png

Downloads & links

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Map

FileDownload / File: emd_9192.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.04 Å
Density
Contour LevelBy AUTHOR: 0.034 / Movie #1: 0.034
Minimum - Maximum-0.05678736 - 0.14083731
Average (Standard dev.)0.00007656962 (±0.0026534924)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 312.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.041.041.04
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z312.000312.000312.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ300300300
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0570.1410.000

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Supplemental data

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Sample components

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Entire : PfRh5/CyRPA/PfRipr complex

EntireName: PfRh5/CyRPA/PfRipr complex
Components
  • Complex: PfRh5/CyRPA/PfRipr complex
    • Protein or peptide: Cysteine-rich protective antigen
    • Protein or peptide: Reticulocyte binding protein 5
    • Protein or peptide: PfRipr
    • Protein or peptide: PfRipr

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Supramolecule #1: PfRh5/CyRPA/PfRipr complex

SupramoleculeName: PfRh5/CyRPA/PfRipr complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Plasmodium falciparum 3D7 (eukaryote)
Recombinant expressionOrganism: Insect cell expression vector pTIE1 (others)

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Macromolecule #1: Cysteine-rich protective antigen

MacromoleculeName: Cysteine-rich protective antigen / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Plasmodium falciparum (isolate 3D7) (eukaryote) / Strain: isolate 3D7
Molecular weightTheoretical: 39.270195 KDa
Recombinant expressionOrganism: Insect cell expression vector pTIE1 (others)
SequenceString: RHVFIRTELS FIKNNVPCIR DMFFIYKREL YNICLDDLKG EEDETHIYVQ KKVKDSWITL NDLFKETDLT GRPHIFAYVD VEEIIILLC EDEEFSNRKK DMTCHRFYSN DGKEYNNAEI TISDYILKDK LLSSYVSLPL KIENREYFLI CGVSPYKFKD D NKKDDILC ...String:
RHVFIRTELS FIKNNVPCIR DMFFIYKREL YNICLDDLKG EEDETHIYVQ KKVKDSWITL NDLFKETDLT GRPHIFAYVD VEEIIILLC EDEEFSNRKK DMTCHRFYSN DGKEYNNAEI TISDYILKDK LLSSYVSLPL KIENREYFLI CGVSPYKFKD D NKKDDILC MASHDKGETW GTKIVIKYDN YKLGVQYFFL RPYISKNDLS FHFYVGDNIN NVKNVNFIEC THEKDLEFVC SN RDFLKDN KVLQDVSTLN DEYIVSYGND NNFAECYIFF NNENSILIKP EKYGNTTAGC YGGTFVKIDE NRALFIYSSS QGI YNIHTI YYANYE

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Macromolecule #2: Reticulocyte binding protein 5

MacromoleculeName: Reticulocyte binding protein 5 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Plasmodium falciparum (malaria parasite P. falciparum)
Molecular weightTheoretical: 39.803371 KDa
Recombinant expressionOrganism: Insect cell expression vector pTIE1 (others)
SequenceString: IIPHYTFLDY YKHLSYNSIY HKSSTYGKCI AVDAFIKKIN ETYDKVKSKC NDIKNDLIAT IKKLEHPYDI NNKNDDSYRY DISEEIDDK SEETDDETEE VEDSIQDTDS NHTPSNKKKN DLMNRTFKKM MDEYNTKKKK LIKCIKNHEN DFNKICMDMK N YGTNLFEQ ...String:
IIPHYTFLDY YKHLSYNSIY HKSSTYGKCI AVDAFIKKIN ETYDKVKSKC NDIKNDLIAT IKKLEHPYDI NNKNDDSYRY DISEEIDDK SEETDDETEE VEDSIQDTDS NHTPSNKKKN DLMNRTFKKM MDEYNTKKKK LIKCIKNHEN DFNKICMDMK N YGTNLFEQ LSCYNNNFCN TNGIRYHYDE YIHKLILSVK SKNLNKDLSD MTNILQQSEL LLTNLNKKMG SYIYIDTIKF IH KEMKHIF NRIEYHTKII NDKTKIIQDK IKLNIWRTFQ KDELLKRILD MSNEYSLFIT SDHLRQMLYN TFYSKEKHLN NIF HHLIYV LQMK

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Macromolecule #3: PfRipr

MacromoleculeName: PfRipr / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Plasmodium falciparum 3D7 (eukaryote)
Molecular weightTheoretical: 1.379692 KDa
Recombinant expressionOrganism: Drosophila nebulosa (fry)
SequenceString:
(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)

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Macromolecule #4: PfRipr

MacromoleculeName: PfRipr / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Plasmodium falciparum 3D7 (eukaryote)
Molecular weightTheoretical: 443.539 Da
Recombinant expressionOrganism: Drosophila nebulosa (fry)
SequenceString:
(UNK)(UNK)(UNK)(UNK)(UNK)

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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

BufferpH: 8.5 / Details: 20 mM Tris, pH 8.5, 150 mM NaCl
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 400
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated magnification: 48077 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 0.01 mm / Nominal magnification: 105000
Specialist opticsSpherical aberration corrector: Microscope was equipped with a Cs corrector with two hexapole elements
Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Sampling interval: 5.0 µm / Number grids imaged: 4 / Number real images: 12974 / Average exposure time: 10.0 sec. / Average electron dose: 92.5 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: Ab initial
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 7.17 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 218837
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model(PDB ID:

4wat

,

5tik

)
RefinementSpace: REAL
Output model

PDB-6mpv:
Cryo-electron microscopy structure of Plasmodium falciparum Rh5/CyRPA/Ripr invasion complex

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