Journal: PLoS Pathog / Year: 2020 Title: Structural and functional evaluation of de novo-designed, two-component nanoparticle carriers for HIV Env trimer immunogens. Authors: Aleksandar Antanasijevic / George Ueda / Philip J M Brouwer / Jeffrey Copps / Deli Huang / Joel D Allen / Christopher A Cottrell / Anila Yasmeen / Leigh M Sewall / Ilja Bontjer / Thomas J ...Authors: Aleksandar Antanasijevic / George Ueda / Philip J M Brouwer / Jeffrey Copps / Deli Huang / Joel D Allen / Christopher A Cottrell / Anila Yasmeen / Leigh M Sewall / Ilja Bontjer / Thomas J Ketas / Hannah L Turner / Zachary T Berndsen / David C Montefiori / Per Johan Klasse / Max Crispin / David Nemazee / John P Moore / Rogier W Sanders / Neil P King / David Baker / Andrew B Ward / Abstract: Two-component, self-assembling nanoparticles represent a versatile platform for multivalent presentation of viral antigens. Computational design of protein nanoparticles with differing sizes and ...Two-component, self-assembling nanoparticles represent a versatile platform for multivalent presentation of viral antigens. Computational design of protein nanoparticles with differing sizes and geometries enables combination with antigens of choice to test novel multimerization concepts in immunization strategies where the goal is to improve the induction and maturation of neutralizing antibody lineages. Here, we describe detailed antigenic, structural, and functional characterization of computationally designed tetrahedral, octahedral, and icosahedral nanoparticle immunogens displaying trimeric HIV envelope glycoprotein (Env) ectodomains. Env trimers, based on subtype A (BG505) or consensus group M (ConM) sequences and engineered with SOSIP stabilizing mutations, were fused to an underlying trimeric building block of each nanoparticle. Initial screening yielded one icosahedral and two tetrahedral nanoparticle candidates, capable of presenting twenty or four copies of the Env trimer. A number of analyses, including detailed structural characterization by cryo-EM, demonstrated that the nanoparticle immunogens possessed the intended structural and antigenic properties. When the immunogenicity of ConM-SOSIP trimers presented on a two-component tetrahedral nanoparticle or as soluble proteins were compared in rabbits, the two immunogens elicited similar serum antibody binding titers against the trimer component. Neutralizing antibody titers were slightly elevated in the animals given the nanoparticle immunogen and were initially more focused to the trimer apex. Altogether, our findings indicate that tetrahedral nanoparticles can be successfully applied for presentation of HIV Env trimer immunogens; however, the optimal implementation to different immunization strategies remains to be determined.
History
Deposition
Jan 5, 2020
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Header (metadata) release
Jan 29, 2020
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Map release
Jul 22, 2020
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Update
Aug 26, 2020
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Current status
Aug 26, 2020
Processing site: RCSB / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Download / File: emd_21180.map.gz / Format: CCP4 / Size: 27 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation
EM-Based Polyclonal Epitope Mapping. ConM-SOSIP Complexed with Purified Polyclonal Fab (Grp 1, ID r2382 Wk22), Negative Stain EM map
Voxel size
X=Y=Z: 2.05 Å
Density
Contour Level
By AUTHOR: 0.02 / Movie #1: 0.02
Minimum - Maximum
-0.02698165 - 0.12725957
Average (Standard dev.)
0.00025611496 (±0.0047337958)
Symmetry
Space group: 1
Details
EMDB XML:
Map geometry
Axis order
X
Y
Z
Origin
0
0
0
Dimensions
192
192
192
Spacing
192
192
192
Cell
A=B=C: 393.59998 Å α=β=γ: 90.0 °
CCP4 map header:
mode
Image stored as Reals
Å/pix. X/Y/Z
2.05
2.05
2.05
M x/y/z
192
192
192
origin x/y/z
0.000
0.000
0.000
length x/y/z
393.600
393.600
393.600
α/β/γ
90.000
90.000
90.000
MAP C/R/S
1
2
3
start NC/NR/NS
0
0
0
NC/NR/NS
192
192
192
D min/max/mean
-0.027
0.127
0.000
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Supplemental data
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Sample components
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Entire : ConM-SOSIP.v9 complexed with purified polyclonal Fab from an immu...
Entire
Name: ConM-SOSIP.v9 complexed with purified polyclonal Fab from an immunized rabbit (Grp 1, ID r2382, Wk22)
Components
Complex: ConM-SOSIP.v9 complexed with purified polyclonal Fab from an immunized rabbit (Grp 1, ID r2382, Wk22)
Protein or peptide: ConM-SOSIP.v9
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Supramolecule #1: ConM-SOSIP.v9 complexed with purified polyclonal Fab from an immu...
Supramolecule
Name: ConM-SOSIP.v9 complexed with purified polyclonal Fab from an immunized rabbit (Grp 1, ID r2382, Wk22) type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Type: NEGATIVE / Material: Uranyl Formate Details: Sample diluted to 0.05 mg/mL. 3 uL was applied onto the grid, blotted off, and then stained with 2% uranyl formate for 60 seconds.
Grid
Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE
Details
ConM-SOSIP.v9 was complexed with the purified polyclonal Fab overnight. The complex was purified using SEC and concentrated. The complex was then diluted in TBS to 0.05mg/ml to be loaded onto an imaging grid.
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Electron microscopy
Microscope
FEI TECNAI SPIRIT
Image recording
Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Number grids imaged: 1 / Average electron dose: 25.0 e/Å2
Electron beam
Acceleration voltage: 120 kV / Electron source: LAB6
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