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John D.
Clarke
,
Helen M. E.
Duyvesteyn
,
Eva
Perez-Martin
,
Undīne
Latišenko
,
Claudine
Porta
,
Kathleen V.
Humphreys
,
Abigail L.
Hay
,
Jingshan
Ren
,
Elizabeth E.
Fry
,
Erwin
Van Den Born
,
Bryan
Charleston
,
Marie
Bonnet-Di Placido
,
Raymond J.
Owens
,
David I.
Stuart
,
John A.
Hammond
Open Access
Abstract: Foot-and-mouth disease vaccination using inactivated virus is suboptimal, as the icosahedral viral capsids often disassemble into antigenically distinct pentameric units during long-term storage, or exposure to elevated temperature or lowered pH, and thus raise a response that is no longer protective. Furthermore, as foot-and-mouth disease virus (FMDV)’s seven serotypes are antigenically diverse, cross-protection from a single serotype vaccine is limited, and most existing mouse and bovine antibodies and camelid single-domain heavy chain-only antibodies are serotype-specific. For quality control purposes, there is a real need for pan-serotype antibodies that clearly distinguish between pentamer (12S) and protective intact FMDV capsid. To date, few cross-serotype bovine-derived antibodies have been reported in the literature. We identify a bovine antibody with an ultralong CDR-H3, Ab117, whose structural analysis reveals that it binds to a deep, hydrophobic pocket on the interior surface of the capsid via the CDR-H3. Main-chain and hydrophobic interactions provide broad serotype specificity. ELISA analysis confirms that Ab117 is a novel pan-serotype and conformational epitope-specific 12S reagent, suitable for assessing capsid integrity.
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Oct 2024
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Helen M. E.
Duyvesteyn
,
Aiste
Dijokaite-Guraliuc
,
Chang
Liu
,
Piyada
Supasa
,
Barbara
Kronsteiner
,
Katie
Jeffery
,
Lizzie
Stafford
,
Paul
Klenerman
,
Susanna J.
Dunachie
,
Juthathip
Mongkolsapaya
,
Elizabeth
Fry
,
Jingshan
Ren
,
David I.
Stuart
,
Gavin R.
Screaton
Abstract: BA.2.87.1 represents a major shift in the BA.2 lineage of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and is unusual in having two lengthy deletions of polypeptide in the spike (S) protein, one of which removes a beta-strand. Here we investigate its neutralization by a variety of sera from infected and vaccinated individuals and determine its spike (S) ectodomain structure. The BA.2.87.1 receptor binding domain (RBD) is structurally conserved and the RBDs are tightly packed in an “all-down” conformation with a small rotation relative to the trimer axis as compared to the closest previously observed conformation. The N-terminal domain (NTD) maintains a remarkably similar structure overall; however, the rearrangements resulting from the deletions essentially destroy the so-called supersite epitope and eliminate one glycan site, while a mutation creates an additional glycan site, effectively shielding another NTD epitope. BA.2.87.1 is relatively easily neutralized but acquisition of additional mutations in the RBD could increase antibody escape allowing it to become a dominant sub-lineage.
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Aug 2024
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I04-Macromolecular Crystallography
VMXi-Versatile Macromolecular Crystallography in situ
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Diamond Proposal Number(s):
[19946, 23570]
Open Access
Abstract: The third complementary-determining regions of the heavy-chain (CDR3H) variable regions (VH) of some cattle antibodies are highly extended, consisting of 48 or more residues. These `ultralong' CDR3Hs form β-ribbon stalks that protrude from the surface of the antibody with a disulfide cross-linked knob region at their apex that dominates antigen interactions over the other CDR loops. The structure of the Fab fragment of a naturally paired bovine ultralong antibody (D08), identified by single B-cell sequencing, has been determined to 1.6 Å resolution. By swapping the D08 native light chain with that of an unrelated antigen-unknown ultralong antibody, it is shown that interactions between the CDR3s of the variable domains potentially affect the fine positioning of the ultralong CDR3H; however, comparison with other crystallographic structures shows that crystalline packing is also a major contributor. It is concluded that, on balance, the exact positioning of ultralong CDR3H loops is most likely to be due to the constraints of crystal packing.
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Jul 2024
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I03-Macromolecular Crystallography
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Gareth T.
Powell
,
Ana
Faro
,
Yuguang
Zhao
,
Heather
Stickney
,
Laura
Novellasdemunt
,
Pedro
Henriques
,
Gaia
Gestri
,
Esther
Redhouse White
,
Jingshan
Ren
,
Weixian
Lu
,
Rodrigo M.
Young
,
Thomas A.
Hawkins
,
Florencia
Cavodeassi
,
Quenten
Schwarz
,
Elena
Dreosti
,
David W.
Raible
,
Vivian S. W.
Li
,
Gavin J.
Wright
,
E. Yvonne
Jones
,
Stephen W.
Wilson
Diamond Proposal Number(s):
[19946]
Abstract: Neurons on the left and right sides of the nervous system often show asymmetric properties, but how such differences arise is poorly understood. Genetic screening in zebrafish revealed that loss of function of the transmembrane protein Cachd1 resulted in right-sided habenula neurons adopting left-sided identity. Cachd1 is expressed in neuronal progenitors, functions downstream of asymmetric environmental signals, and influences timing of the normally asymmetric patterns of neurogenesis. Biochemical and structural analyses demonstrated that Cachd1 can bind simultaneously to Lrp6 and Frizzled family Wnt co-receptors. Consistent with this, lrp6 mutant zebrafish lose asymmetry in the habenulae, and epistasis experiments support a role for Cachd1 in modulating Wnt pathway activity in the brain. These studies identify Cachd1 as a conserved Wnt receptor–interacting protein that regulates lateralized neuronal identity in the zebrafish brain.
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May 2024
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I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Chang
Liu
,
Raksha
Das
,
Aiste
Dijokaite-Guraliuc
,
Daming
Zhou
,
Alexander J.
Mentzer
,
Piyada
Supasa
,
Muneeswaran
Selvaraj
,
Helen M. E.
Duyvesteyn
,
Thomas G.
Ritter
,
Nigel
Temperton
,
Paul
Klenerman
,
Susanna J.
Dunachie
,
Neil G.
Paterson
,
Mark A.
Williams
,
David R.
Hall
,
Elizabeth E.
Fry
,
Juthathip
Mongkolsapaya
,
Jingshan
Ren
,
David I.
Stuart
,
Gavin R.
Screaton
Open Access
Abstract: The rapid evolution of SARS-CoV-2 is driven in part by a need to evade the antibody response in the face of high levels of immunity. Here, we isolate spike (S) binding monoclonal antibodies (mAbs) from vaccinees who suffered vaccine break-through infections with Omicron sub lineages BA.4 or BA.5. Twenty eight potent antibodies are isolated and characterised functionally, and in some cases structurally. Since the emergence of BA.4/5, SARS-CoV-2 has continued to accrue mutations in the S protein, to understand this we characterize neutralization of a large panel of variants and demonstrate a steady attrition of neutralization by the panel of BA.4/5 mAbs culminating in total loss of function with recent XBB.1.5.70 variants containing the so-called ‘FLip’ mutations at positions 455 and 456. Interestingly, activity of some mAbs is regained on the recently reported variant BA.2.86.
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Apr 2024
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Daming
Zhou
,
Piyada
Supasa
,
Chang
Liu
,
Aiste
Dijokaite-Guraliuc
,
Helen M. E.
Duyvesteyn
,
Muneeswaran
Selvaraj
,
Alexander J.
Mentzer
,
Raksha
Das
,
Wanwisa
Dejnirattisai
,
Nigel
Temperton
,
Paul
Klenerman
,
Susanna J.
Dunachie
,
Elizabeth E.
Fry
,
Juthathip
Mongkolsapaya
,
Jingshan
Ren
,
David I.
Stuart
,
Gavin R.
Screaton
Open Access
Abstract: Under pressure from neutralising antibodies induced by vaccination or infection the SARS-CoV-2 spike gene has become a hotspot for evolutionary change, leading to the failure of all mAbs developed for clinical use. Most potent antibodies bind to the receptor binding domain which has become heavily mutated. Here we study responses to a conserved epitope in sub-domain-1 (SD1) of spike which have become more prominent because of mutational escape from antibodies directed to the receptor binding domain. Some SD1 reactive mAbs show potent and broad neutralization of SARS-CoV-2 variants. We structurally map the dominant SD1 epitope and provide a mechanism of action by blocking interaction with ACE2. Mutations in SD1 have not been sustained to date, but one, E554K, leads to escape from mAbs. This mutation has now emerged in several sublineages including BA.2.86, reflecting selection pressure on the virus exerted by the increasing prominence of the anti-SD1 response.
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Mar 2024
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Open Access
Abstract: The COVID-19 pandemic caused by SARS-CoV-2 has led to hundreds of millions of infections and millions of deaths, however, human monoclonal antibodies (mAbs) can be an effective treatment. Since SARS-CoV-2 emerged, a variety of strains have acquired increasing numbers of mutations to gain increased transmissibility and escape from the immune response. Most reported neutralizing human mAbs, including all approved therapeutic ones, have been knocked down or out by these mutations. Broadly neutralizing mAbs are therefore of great value, to treat current and possible future variants. Here, we review four types of neutralizing mAbs against the spike protein with broad potency against previously and currently circulating variants. These mAbs target the receptor-binding domain, the subdomain 1, the stem helix, or the fusion peptide. Understanding how these mAbs retain potency in the face of mutational change could guide future development of therapeutic antibodies and vaccines.
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Aug 2023
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I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Aiste
Dijokaite-Guraliuc
,
Raksha
Das
,
Daming
Zhou
,
Helen M.
Ginn
,
Chang
Liu
,
Helen M. E.
Duyvesteyn
,
Jiandong
Huo
,
Rungtiwa
Nutalai
,
Piyada
Supasa
,
Muneeswaran
Selvaraj
,
Thushan I.
De Silva
,
Megan
Plowright
,
Thomas A. H.
Newman
,
Hailey
Hornsby
,
Alexander J.
Mentzer
,
Donal
Skelly
,
Thomas G.
Ritter
,
Nigel
Temperton
,
Paul
Klenerman
,
Eleanor
Barnes
,
Susanna J.
Dunachie
,
Cornelius
Roemer
,
Thomas P.
Peacock
,
Neil G.
Paterson
,
Mark A.
Williams
,
David R.
Hall
,
Elizabeth E.
Fry
,
Juthathip
Mongkolsapaya
,
Jingshan
Ren
,
David I.
Stuart
,
Gavin R.
Screaton
Diamond Proposal Number(s):
[27009]
Open Access
Abstract: In November 2021 Omicron BA.1, containing a raft of new spike mutations emerged and quickly spread globally. Intense selection pressure to escape the antibody response produced by vaccines or SARS-CoV-2 infection then led to a rapid succession of Omicron sub-lineages with waves of BA.2 then BA.4/5 infection. Recently, many variants have emerged such as BQ.1 and XBB, which carry up to 8 additional RBD amino-acid substitutions compared to BA.2. We describe a panel of 25 potent mAbs generated from vaccinees suffering BA.2 breakthrough infections. Epitope mapping shows potent mAb binding shifting to 3 clusters, 2 corresponding to early-pandemic binding hotspots. The RBD mutations in recent variants map close to these binding sites and knock out or severely knock down neutralization activity of all but 1 potent mAb. This recent mAb escape corresponds with large falls in neutralization titre of vaccine or BA.1, BA.2 or BA.4/5 immune serum.
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Mar 2023
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I03-Macromolecular Crystallography
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Jiandong
Huo
,
Aiste
Dijokaite-Guraliuc
,
Chang
Liu
,
Raksha
Das
,
Piyada
Supasa
,
Muneeswaran
Selvaraj
,
Rungtiwa
Nutalai
,
Daming
Zhou
,
Alexander J.
Mentzer
,
Donal
Skelly
,
Thomas G.
Ritter
,
Ali
Amini
,
Sagida
Bibi
,
Sandra
Adele
,
Sile Ann
Johnson
,
Neil G.
Paterson
,
Mark A.
Williams
,
David R.
Hall
,
Megan
Plowright
,
Thomas A. H.
Newman
,
Hailey
Hornsby
,
Thushan I.
De Silva
,
Nigel
Temperton
,
Paul
Klenerman
,
Eleanor
Barnes
,
Susanna J.
Dunachie
,
Andrew J.
Pollard
,
Teresa
Lambe
,
Philip
Goulder
,
Elizabeth E.
Fry
,
Juthathip
Mongkolsapaya
,
Jingshan
Ren
,
David I.
Stuart
,
Gavin R.
Screaton
Diamond Proposal Number(s):
[27009]
Open Access
Abstract: Variants of SARS CoV-2 have caused successive global waves of infection. These variants, with multiple mutations in the spike protein are thought to facilitate escape from natural and vaccine-induced immunity and often increase in the affinity for ACE2. The latest variant to cause concern is BA.2.75, identified in India where it is now the dominant strain, with evidence of wider dissemination. BA.2.75 is derived from BA.2 and contains four additional mutations in the receptor binding domain (RBD). Here we perform an antigenic and biophysical characterization of BA.2.75, revealing an interesting balance between humoral evasion and ACE2 receptor affinity. ACE2 affinity for BA.2.75 is increased 9-fold compared to BA.2; there is also evidence of escape of BA.2.75 from immune serum, particularly that induced by Delta infection which may explain the rapid spread in India, where BA.2.75 is now the dominant variant. ACE2 affinity appears to be prioritised over greater escape.
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Dec 2022
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Open Access
Abstract: Echoviruses, for which there are currently no approved vaccines or drugs, are responsible for a range of human diseases, for example echovirus 11 (E11) is a major cause of serious neonatal morbidity and mortality. Decay-accelerating factor (DAF, also known as CD55) is an attachment receptor for E11. Here, we report the structure of the complex of E11 and the full-length ectodomain of DAF (short consensus repeats, SCRs, 1–4) at 3.1 Å determined by cryo-electron microscopy (cryo-EM). SCRs 3 and 4 of DAF interact with E11 at the southern rim of the canyon via the VP2 EF and VP3 BC loops. We also observe an unexpected interaction between the N-linked glycan (residue 95 of DAF) and the VP2 BC loop of E11. DAF is a receptor for at least 20 enteroviruses and we classify its binding patterns from reported DAF/virus complexes into two distinct positions and orientations, named as E6 and E11 poses. Whilst 60 DAF molecules can attach to the virion in the E6 pose, no more than 30 can attach to E11 due to steric restrictions. Analysis of the distinct modes of interaction and structure and sequence-based phylogenies suggests that the two modes evolved independently, with the E6 mode likely found earlier.
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Nov 2022
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