Structural basis of Omicron neutralization by affinity-matured public antibodies

The SARS-CoV-2 Omicron Variant of Concern (B.1.1.529) has spread rapidly in many countries. With a spike that is highly diverged from that of the pandemic founder, it escapes most available monoclonal antibody therapeutics and erodes vaccine protection. A public class of IGHV3-53-using SARS-CoV-2 neutralizing antibodies typically fails to neutralize variants carrying mutations in the receptor-binding motif, including Omicron. As antibodies from this class are likely elicited in most people following SARS-CoV-2 infection or vaccination, their subsequent affinity maturation is of particular interest. Here, we isolated IGHV3-53-using antibodies from an individual seven months after infection and identified several antibodies capable of broad and potent SARS-CoV-2 neutralization, extending to Omicron without loss of potency. By introducing select somatic hypermutations into a germline-reverted form of one such antibody, CAB-A17, we demonstrate the potential for commonly elicited antibodies to develop broad cross-neutralization through affinity maturation. Further, we resolved the structure of CAB-A17 Fab in complex with Omicron spike at an overall resolution of 2.6 angstroms by cryo-electron microscopy and defined the structural basis for this breadth. Thus, public SARS-CoV-2 neutralizing antibodies can, without modified spike vaccines, mature to cross-neutralize exceptionally antigenically diverged SARS-CoV-2 variants.

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Structural basis of SARS-CoV-2 Omicron immune evasion and receptor engagement

10.1101/2021.12.28.474380 ◽

2021 ◽

Author(s):

Matthew McCallum ◽

Nadine Czudnochowski ◽

Laura E Rosen ◽

Samantha K Zepeda ◽

John E Bowen ◽

...

Keyword(s):

Electron Microscopy ◽

Monoclonal Antibody ◽

Crystal Structures ◽

Immune Evasion ◽

Marked Reduction ◽

Structural Basis ◽

X Ray ◽

Structural Framework ◽

Cryo Electron Microscopy ◽

Antigenic Shift

The SARS-CoV-2 Omicron variant of concern evades antibody mediated immunity with an unprecedented magnitude due to accumulation of numerous spike mutations. To understand the Omicron antigenic shift, we determined cryo-electron microscopy and X-ray crystal structures of the spike and RBD bound to the broadly neutralizing sarbecovirus monoclonal antibody (mAb) S309 (the parent mAb of sotrovimab) and to the human ACE2 receptor. We provide a structural framework for understanding the marked reduction of binding of all other therapeutic mAbs leading to dampened neutralizing activity. We reveal electrostatic remodeling of the interactions within the spike and those formed between the Omicron RBD and human ACE2, likely explaining enhanced affinity for the host receptor relative to the prototypic virus.

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Structural basis for potent neutralization of SARS-CoV-2 and role of antibody affinity maturation

Nature Communications ◽

10.1038/s41467-020-19231-9 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 4

Author(s):

Nicholas K. Hurlburt ◽

Emilie Seydoux ◽

Yu-Hsin Wan ◽

Venkata Viswanadh Edara ◽

Andrew B. Stuart ◽

...

Keyword(s):

Receptor Binding ◽

Neutralizing Antibodies ◽

Substantial Reduction ◽

Affinity Maturation ◽

Structural Basis ◽

Binding Motif ◽

Neutralization Potential ◽

Neutralizing Monoclonal Antibody ◽

Potent Neutralization

Abstract SARS-CoV-2 is a betacoronavirus virus responsible for the COVID-19 pandemic. Here, we determine the X-ray crystal structure of a potent neutralizing monoclonal antibody, CV30, isolated from a patient infected with SARS-CoV-2, in complex with the receptor binding domain. The structure reveals that CV30 binds to an epitope that overlaps with the human ACE2 receptor binding motif providing a structural basis for its neutralization. CV30 also induces shedding of the S1 subunit, indicating an additional mechanism of neutralization. A germline reversion of CV30 results in a substantial reduction in both binding affinity and neutralization potential indicating the minimal somatic mutation is needed for potently neutralizing antibodies against SARS-CoV-2.

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Structural basis for antibody resistance to SARS-CoV-2 omicron variant

10.1101/2021.12.21.473620 ◽

2021 ◽

Author(s):

Gabriele Cerutti ◽

Yicheng Guo ◽

Liu Lihong ◽

Zhening Zhang ◽

Liyuan Liu ◽

...

Keyword(s):

Electron Microscopy ◽

Receptor Binding ◽

Neutralizing Antibodies ◽

Spike Protein ◽

Structural Basis ◽

Receptor Binding Domain ◽

Functional Studies ◽

Antibody Recognition ◽

Cryo Electron Microscopy ◽

Antibody Resistance

The recently reported B.1.1.529 Omicron variant of SARS-CoV-2 includes 34 mutations in the spike protein relative to the Wuhan strain that initiated the COVID-19 pandemic, including 15 mutations in the receptor binding domain (RBD). Functional studies have shown omicron to substantially escape the activity of many SARS-CoV-2-neutralizing antibodies. Here we report a 3.1 Å resolution cryo-electron microscopy (cryo-EM) structure of the Omicron spike protein ectodomain. The structure depicts a spike that is exclusively in the 1-RBD-up conformation with increased mobility and inter-protomer asymmetry. Many mutations cause steric clashes and/or altered interactions at antibody binding surfaces, whereas others mediate changes of the spike structure in local regions to interfere with antibody recognition. Overall, the structure of the omicron spike reveals how mutations alter its conformation and explains its extraordinary ability to evade neutralizing antibodies.

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A broadly neutralizing macaque monoclonal antibody against the HIV-1 V3-Glycan patch

eLife ◽

10.7554/elife.61991 ◽

2020 ◽

Vol 9 ◽

Cited By ~ 1

Author(s):

Zijun Wang ◽

Christopher O Barnes ◽

Rajeev Gautam ◽

Julio C Cetrulo Lorenzi ◽

Christian T Mayer ◽

...

Keyword(s):

Electron Microscopy ◽

Monoclonal Antibody ◽

Neutralizing Antibodies ◽

Geometric Mean ◽

High Dose ◽

Dependent Manner ◽

Peptide Motif ◽

Broadly Neutralizing Antibodies ◽

Cryo Electron Microscopy ◽

Hiv 1

A small fraction of HIV-1- infected humans develop broadly neutralizing antibodies (bNAbs) against HIV-1 that protect macaques from simian immunodeficiency HIV chimeric virus (SHIV). Similarly, a small number of macaques infected with SHIVs develop broadly neutralizing serologic activity, but less is known about the nature of simian antibodies. Here, we report on a monoclonal antibody, Ab1485, isolated from a macaque infected with SHIVAD8 that developed broadly neutralizing serologic activity targeting the V3-glycan region of HIV-1 Env. Ab1485 neutralizes 38.1% of HIV-1 isolates in a 42-pseudovirus panel with a geometric mean IC50 of 0.055 µg/mLl and SHIVAD8 with an IC50 of 0.028 µg/mLl. Ab1485 binds the V3-glycan epitope in a glycan-dependent manner. A 3.5 Å cryo-electron microscopy structure of Ab1485 in complex with a native-like SOSIP Env trimer showed conserved contacts with the N332gp120 glycan and gp120 GDIR peptide motif, but in a distinct Env-binding orientation relative to human V3/N332gp120 glycan-targeting bNAbs. Intravenous infusion of Ab1485 protected macaques from a high dose challenge with SHIVAD8. We conclude that macaques can develop bNAbs against the V3-glycan patch that resemble human V3-glycan bNAbs.

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Structural basis of a potent human monoclonal antibody against Zika virus targeting a quaternary epitope

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1815432116 ◽

2019 ◽

Vol 116 (5) ◽

pp. 1591-1596 ◽

Cited By ~ 26

Author(s):

Feng Long ◽

Michael Doyle ◽

Estefania Fernandez ◽

Andrew S. Miller ◽

Thomas Klose ◽

...

Keyword(s):

Monoclonal Antibody ◽

Zika Virus ◽

Neutralizing Antibodies ◽

Human Monoclonal Antibody ◽

Structural Basis ◽

E Proteins ◽

Fab Fragment ◽

Cryo Electron Microscopy ◽

E Protein ◽

Mouse Model Of Infection

Zika virus (ZIKV) is a major human pathogen and member of the Flavivirus genus in the Flaviviridae family. In contrast to most other insect-transmitted flaviviruses, ZIKV also can be transmitted sexually and from mother to fetus in humans. During recent outbreaks, ZIKV infections have been linked to microcephaly, congenital disease, and Guillain-Barré syndrome. Neutralizing antibodies have potential as therapeutic agents. We report here a 4-Å-resolution cryo-electron microscopy structure of the ZIKV virion in complex with Fab fragments of the potently neutralizing human monoclonal antibody ZIKV-195. The footprint of the ZIKV-195 Fab fragment expands across two adjacent envelope (E) protein protomers. ZIKV neutralization by this antibody is presumably accomplished by cross-linking the E proteins, which likely prevents formation of E protein trimers required for fusion of the viral and cellular membranes. A single dose of ZIKV-195 administered 5 days after virus inoculation showed marked protection against lethality in a stringent mouse model of infection.

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A 2.8-Angstrom-Resolution Cryo-Electron Microscopy Structure of Human Parechovirus 3 in Complex with Fab from a Neutralizing Antibody

Journal of Virology ◽

10.1128/jvi.01597-18 ◽

2018 ◽

Vol 93 (4) ◽

Cited By ~ 5

Author(s):

Aušra Domanska ◽

Justin W. Flatt ◽

Joonas J. J. Jukonen ◽

James A. Geraets ◽

Sarah J. Butcher

Keyword(s):

Electron Microscopy ◽

Monoclonal Antibody ◽

High Resolution ◽

Cultured Cells ◽

Human Monoclonal Antibody ◽

Neutralizing Antibody ◽

Molecular Diagnostic ◽

Diagnostic Tools ◽

Human Parechovirus ◽

Cryo Electron Microscopy

ABSTRACTHuman parechovirus 3 (HPeV3) infection is associated with sepsis characterized by significant immune activation and subsequent tissue damage in neonates. Strategies to limit infection have been unsuccessful due to inadequate molecular diagnostic tools for early detection and the lack of a vaccine or specific antiviral therapy. Toward the latter, we present a 2.8-Å-resolution structure of HPeV3 in complex with fragments from a neutralizing human monoclonal antibody, AT12-015, using cryo-electron microscopy (cryo-EM) and image reconstruction. Modeling revealed that the epitope extends across neighboring asymmetric units with contributions from capsid proteins VP0, VP1, and VP3. Antibody decoration was found to block binding of HPeV3 to cultured cells. Additionally, at high resolution, it was possible to model a stretch of RNA inside the virion and, from this, identify the key features that drive and stabilize protein-RNA association during assembly.IMPORTANCEHuman parechovirus 3 (HPeV3) is receiving increasing attention as a prevalent cause of sepsis-like symptoms in neonates, for which, despite the severity of disease, there are no effective treatments available. Structural and molecular insights into virus neutralization are urgently needed, especially as clinical cases are on the rise. Toward this goal, we present the first structure of HPeV3 in complex with fragments from a neutralizing monoclonal antibody. At high resolution, it was possible to precisely define the epitope that, when targeted, prevents virions from binding to cells. Such an atomic-level description is useful for understanding host-pathogen interactions and viral pathogenesis mechanisms and for finding potential cures for infection and disease.

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The U4 Antibody Epitope on Human Papillomavirus 16 Identified by Cryo-electron Microscopy

Journal of Virology ◽

10.1128/jvi.02020-15 ◽

2015 ◽

Vol 89 (23) ◽

pp. 12108-12117 ◽

Cited By ~ 16

Author(s):

Jian Guan ◽

Stephanie M. Bywaters ◽

Sarah A. Brendle ◽

Hyunwook Lee ◽

Robert E. Ashley ◽

...

Keyword(s):

Electron Microscopy ◽

Monoclonal Antibody ◽

Human Papillomavirus ◽

Conformational Changes ◽

Vaccine Development ◽

Contact Point ◽

Human Papillomavirus 16 ◽

Cryo Electron Microscopy ◽

C Terminus ◽

L1 Protein

ABSTRACTThe human papillomavirus (HPV) major structural protein L1 composes capsomers that are linked together through interactions mediated by the L1 C terminus to constitute a T=7 icosahedral capsid. H16.U4 is a type-specific monoclonal antibody recognizing a conformation-dependent neutralizing epitope of HPV thought to include the L1 protein C terminus. The structure of human papillomavirus 16 (HPV16) complexed with H16.U4 fragments of antibody (Fab) was solved by cryo-electron microscopy (cryo-EM) image reconstruction. Atomic structures of virus and Fab were fitted into the corresponding cryo-EM densities to identify the antigenic epitope. The antibody footprint mapped predominately to the L1 C-terminal arm with an additional contact point on the side of the capsomer. This footprint describes an epitope that is presented capsid-wide. However, although the H16.U4 epitope suggests the presence of 360 potential binding sites exposed in the capsid valley between each capsomer, H16.U4 Fab bound only to epitopes located around the icosahedral five-fold vertex of the capsid. Thus, the binding characteristics of H16.U4 defined in this study showed a distinctive selectivity for local conformation-dependent interactions with specific L1 invading arms between five-fold related capsomers.IMPORTANCEHuman papillomavirus 16 (HPV16) is the most prevalent oncogenic genotype in HPV-associated anogenital and oral cancers. Here we use cryo-EM reconstruction techniques to solve the structures of the HPV16 capsid complexes using H16.U4 fragment of antibody (Fab). Different from most other antibodies directed against surface loops, H16.U4 monoclonal antibody is unique in targeting the C-terminal arm of the L1 protein. This monoclonal antibody (MAb) is used throughout the HPV research community in HPV serological and vaccine development and to define mechanisms of HPV uptake. The unique binding mode of H16.U4 defined here shows important conformation-dependent interactions within the HPV16 capsid. By targeting an important structural and conformational epitope, H16.U4 may identify subtle conformational changes in different maturation stages of the HPV capsid and provide a key probe to analyze the mechanisms of HPV uptake during the early stages of virus infection. Our analyses precisely define important conformational epitopes on HPV16 capsids that are key targets for successful HPV prophylactic vaccines.

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Cryo-EM structure and polymorphism of Aβ amyloid fibrils purified from Alzheimer’s brain tissue

Nature Communications ◽

10.1038/s41467-019-12683-8 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 85

Author(s):

Marius Kollmer ◽

William Close ◽

Leonie Funk ◽

Jay Rasmussen ◽

Aref Bsoul ◽

...

Keyword(s):

Electron Microscopy ◽

Brain Tissue ◽

Amyloid Fibrils ◽

Structural Basis ◽

Amyloid Angiopathy ◽

Cryo Electron Microscopy ◽

Aβ Amyloid ◽

Cerebral Amyloid ◽

Aβ Fibrils

Abstract The formation of Aβ amyloid fibrils is a neuropathological hallmark of Alzheimer’s disease and cerebral amyloid angiopathy. However, the structure of Aβ amyloid fibrils from brain tissue is poorly understood. Here we report the purification of Aβ amyloid fibrils from meningeal Alzheimer’s brain tissue and their structural analysis with cryo-electron microscopy. We show that these fibrils are polymorphic but consist of similarly structured protofilaments. Brain derived Aβ amyloid fibrils are right-hand twisted and their peptide fold differs sharply from previously analyzed Aβ fibrils that were formed in vitro. These data underscore the importance to use patient-derived amyloid fibrils when investigating the structural basis of the disease.

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How Good Can Single-Particle Cryo-EM Become? What Remains Before It Approaches Its Physical Limits?

Annual Review of Biophysics ◽

10.1146/annurev-biophys-070317-032828 ◽

2019 ◽

Vol 48 (1) ◽

pp. 45-61 ◽

Cited By ~ 26

Author(s):

Robert M. Glaeser

Keyword(s):

Electron Microscopy ◽

High Resolution ◽

Single Particle ◽

Energy Use ◽

Optimal Choice ◽

Structural Basis ◽

Detective Quantum Efficiency ◽

Quantum Metrology ◽

Cryo Electron Microscopy ◽

Induced Motion

Impressive though the achievements of single-particle cryo–electron microscopy are today, a substantial gap still remains between what is currently accomplished and what is theoretically possible. As is reviewed here, twofold or more improvements are possible as regards ( a) the detective quantum efficiency of cameras at high resolution, ( b) converting phase modulations to intensity modulations in the image, and ( c) recovering the full amount of high-resolution signal in the presence of beam-induced motion of the specimen. In addition, potential for improvement is reviewed for other topics such as optimal choice of electron energy, use of aberration correctors, and quantum metrology. With the help of such improvements, it does not seem to be too much to imagine that determining the structural basis for every aspect of catalytic control, signaling, and regulation, in any type of cell of interest, could easily be accelerated fivefold or more.

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