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- EMDB-27222: CIS43 Variant 10 bound to pfCSP Class 3 -

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Basic information

Entry
Database: EMDB / ID: EMD-27222
TitleCIS43 Variant 10 bound to pfCSP Class 3
Map dataUnsharpened map
Sample
  • Complex: CIS43 Variant 10 bound to pfCSP Class 3
    • Protein or peptide: Var10 Light
    • Protein or peptide: Var10 Heavy
    • Protein or peptide: pfCSPm
Biological speciesPlasmodium falciparum (malaria parasite P. falciparum) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 9.8 Å
AuthorsGorman J / Kwong PD
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
Other privateSimons Foundation (SF349247)
CitationJournal: J Exp Med / Year: 2022
Title: Highly protective antimalarial antibodies via precision library generation and yeast display screening.
Authors: Bailey B Banach / Prabhanshu Tripathi / Lais Da Silva Pereira / Jason Gorman / Thuy Duong Nguyen / Marlon Dillon / Ahmed S Fahad / Patience K Kiyuka / Bharat Madan / Jacy R Wolfe / Brian ...Authors: Bailey B Banach / Prabhanshu Tripathi / Lais Da Silva Pereira / Jason Gorman / Thuy Duong Nguyen / Marlon Dillon / Ahmed S Fahad / Patience K Kiyuka / Bharat Madan / Jacy R Wolfe / Brian Bonilla / Barbara Flynn / Joseph R Francica / Nicholas K Hurlburt / Neville K Kisalu / Tracy Liu / Li Ou / Reda Rawi / Arne Schön / Chen-Hsiang Shen / I-Ting Teng / Baoshan Zhang / Marie Pancera / Azza H Idris / Robert A Seder / Peter D Kwong / Brandon J DeKosky /
Abstract: The monoclonal antibody CIS43 targets the Plasmodium falciparum circumsporozoite protein (PfCSP) and prevents malaria infection in humans for up to 9 mo following a single intravenous administration. ...The monoclonal antibody CIS43 targets the Plasmodium falciparum circumsporozoite protein (PfCSP) and prevents malaria infection in humans for up to 9 mo following a single intravenous administration. To enhance the potency and clinical utility of CIS43, we used iterative site-saturation mutagenesis and DNA shuffling to screen precise gene-variant yeast display libraries for improved PfCSP antigen recognition. We identified several mutations that improved recognition, predominately in framework regions, and combined these to produce a panel of antibody variants. The most improved antibody, CIS43_Var10, had three mutations and showed approximately sixfold enhanced protective potency in vivo compared to CIS43. Co-crystal and cryo-electron microscopy structures of CIS43_Var10 with the peptide epitope or with PfCSP, respectively, revealed functional roles for each of these mutations. The unbiased site-directed mutagenesis and screening pipeline described here represent a powerful approach to enhance protective potency and to enable broader clinical use of antimalarial antibodies.
History
DepositionJun 6, 2022-
Header (metadata) releaseJun 29, 2022-
Map releaseJun 29, 2022-
UpdateJul 6, 2022-
Current statusJul 6, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27222.png

Downloads & links

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Map

FileDownload / File: emd_27222.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationUnsharpened map
Voxel sizeX=Y=Z: 1.083 Å
Density
Contour LevelBy AUTHOR: 0.16
Minimum - Maximum-0.20689577 - 0.7048578
Average (Standard dev.)0.00086231646 (±0.024737827)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 346.56 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_27222_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_27222_half_map_1.map
Projections & Slices
AxesZYX

Projections

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_27222_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : CIS43 Variant 10 bound to pfCSP Class 3

EntireName: CIS43 Variant 10 bound to pfCSP Class 3
Components
  • Complex: CIS43 Variant 10 bound to pfCSP Class 3
    • Protein or peptide: Var10 Light
    • Protein or peptide: Var10 Heavy
    • Protein or peptide: pfCSPm

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Supramolecule #1: CIS43 Variant 10 bound to pfCSP Class 3

SupramoleculeName: CIS43 Variant 10 bound to pfCSP Class 3 / type: complex / Chimera: Yes / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Plasmodium falciparum (malaria parasite P. falciparum)

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Macromolecule #1: Var10 Light

MacromoleculeName: Var10 Light / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString: DIVMTQSPDS LAVSLGERAT INCKSSQSVF YSSNNKNYLA WYQQKPGQPP NLLIYWASTR QSGVPDRFSG SGSGTDFTL TISSLQAEDV ASYYCHQYYS SPLTFGGGTK VEIKRTVAAP SVFIFPPSDE QLKSGTASVV C LLNNFYPR EAKVQWKVDN ALQSGNSQES ...String:
DIVMTQSPDS LAVSLGERAT INCKSSQSVF YSSNNKNYLA WYQQKPGQPP NLLIYWASTR QSGVPDRFSG SGSGTDFTL TISSLQAEDV ASYYCHQYYS SPLTFGGGTK VEIKRTVAAP SVFIFPPSDE QLKSGTASVV C LLNNFYPR EAKVQWKVDN ALQSGNSQES VTEQDSKDST YSLSSTLTLS KADYEKHKVY ACEVTHQGLS SP VTKSFNR

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Macromolecule #2: Var10 Heavy

MacromoleculeName: Var10 Heavy / type: protein_or_peptide / ID: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
SequenceString: QRQLVQSGAE VKKPGASVKV SCKASGYTFT SYAIHWVRQA PGQRLEWMGW IKAGNGNTRY SQKFQDRVTI TRDTSTTTA YMELSSLRSE DTAVYYCALL TVLTPDDAFD IWGQGTMVTV SSASTKGPSV FPLAPSSKST S GGTAALGC LVKDYFPEPV TVSWNSGALT ...String:
QRQLVQSGAE VKKPGASVKV SCKASGYTFT SYAIHWVRQA PGQRLEWMGW IKAGNGNTRY SQKFQDRVTI TRDTSTTTA YMELSSLRSE DTAVYYCALL TVLTPDDAFD IWGQGTMVTV SSASTKGPSV FPLAPSSKST S GGTAALGC LVKDYFPEPV TVSWNSGALT SGVHTFPAVL QSSGLYSLSS VVTVPSSSLG TQTYICNVNH KP SNTKVDK KVEP

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Macromolecule #3: pfCSPm

MacromoleculeName: pfCSPm / type: protein_or_peptide / ID: 3 / Enantiomer: LEVO
Source (natural)Organism: Plasmodium falciparum (malaria parasite P. falciparum)
SequenceString: IATMDSKGSS QKGSRLLLLL VVSNLLLPQG VLAQEYQSYG SSSNTRVLNE LNYDNAGTNL YNELEMNYYG KQENWYSLSS NSASLGENDD GNNEDNEKLR KPKHKKLKQP ADGNPDP NA NPNVDPNANP NVDPNANPNV DPNANPNANP NANPNANPNA NPNANPNANP ...String:
IATMDSKGSS QKGSRLLLLL VVSNLLLPQG VLAQEYQSYG SSSNTRVLNE LNYDNAGTNL YNELEMNYYG KQENWYSLSS NSASLGENDD GNNEDNEKLR KPKHKKLKQP ADGNPDP NA NPNVDPNANP NVDPNANPNV DPNANPNANP NANPNANPNA NPNANPNANP NANPNANPNA NPNANPNANP NANPNANPNA NPNANPNANP NVDPNANPNA NPNAN PNAN PNANPNANPN ANPNANPNAN PNANPNANPN ANPNANPNAN PNANPNANPN ANPNANPNKN NQGNGQGHNM PNDPNRNVDE NANANSAVKN NNNEEPSDKH IKEY LNKIQ NSLSTEWSPC SVTCGNGIQV RIKPGSANKP KDELDYANDI EKKICKMEKC SGSGLNDIFE AQKIEWHELE VLFQ

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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

Concentration2 mg/mL
BufferpH: 7.4 / Details: PBS
GridModel: C-flat-1.2/1.3 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV
DetailsVar10 Fab with pfCSPm

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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 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Average exposure time: 2.0 sec. / Average electron dose: 64.12 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: cryoSPARC (ver. 3)
Initial angle assignmentType: NOT APPLICABLE
Final 3D classificationSoftware - Name: cryoSPARC (ver. 3)
Final angle assignmentType: NOT APPLICABLE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 9.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3) / Number images used: 28900

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