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12 items found (0 items not approved), displaying 1 to 10.[First/Prev] 1, 2 [Next/Last]

Instruments / Technical Area	Publication Details	Published
I03-Macromolecular Crystallography	Humoral responses against SARS-CoV-2 Omicron BA.2.11, BA.2.12.1 and BA.2.13 from vaccine and BA.1 serum Jiandong Huo , Aiste Dijokaite-Guraliuc , Rungtiwa Nutalai , Raksha Das , Daming Zhou , Alexander J. Mentzer , Elizabeth E. Fry , Juthathip Mongkolsapaya , Jingshan Ren , David I. Stuart , Gavin R. Screaton Cell Discovery 8 DOI: 10.1038/s41421-022-00482-3 Diamond Proposal Number(s): [27009] Open Access	Nov 2022
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	GPC3-Unc5 receptor complex structure and role in cell migration Onno Akkermans , Céline Delloye-Bourgeois , Claudia Peregrina , Maria Carrasquero-Ordaz , Maria Kokolaki , Miguel Berbeira-Santana , Matthieu Chavent , Florie Reynaud , Ritu Raj , Jon Agirre , Metin Aksu , Eleanor S. White , Edward Lowe , Dounia Ben Amar , Sofia Zaballa , Jiandong Huo , Irene Pakos , Patrick T. N. Mccubbin , Davide Comoletti , Raymond J. Owens , Carol V. Robinson , Valérie Castellani , Daniel Del Toro , Elena Seiradake Cell 185, 3931 - 3949.e26 DOI: 10.1016/j.cell.2022.09.025 Diamond Proposal Number(s): [18069] Open Access Show Abstract ▼ Abstract: Neural migration is a critical step during brain development that requires the interactions of cell-surface guidance receptors. Cancer cells often hijack these mechanisms to disseminate. Here, we reveal crystal structures of Uncoordinated-5 receptor D (Unc5D) in complex with morphogen receptor glypican-3 (GPC3), forming an octameric glycoprotein complex. In the complex, four Unc5D molecules pack into an antiparallel bundle, flanked by four GPC3 molecules. Central glycan-glycan interactions are formed by N-linked glycans emanating from GPC3 (N241 in human) and C-mannosylated tryptophans of the Unc5D thrombospondin-like domains. MD simulations, mass spectrometry and structure-based mutants validate the crystallographic data. Anti-GPC3 nanobodies enhance or weaken Unc5-GPC3 binding and, together with mutant proteins, show that Unc5/GPC3 guide migrating pyramidal neurons in the mouse cortex, and cancer cells in an embryonic xenograft neuroblastoma model. The results demonstrate a conserved structural mechanism of cell guidance, where finely balanced Unc5-GPC3 interactions regulate cell migration.	Oct 2022
I03-Macromolecular Crystallography Krios II-Titan Krios II at Diamond Krios IV-Titan Krios IV at Diamond	Correlation between the binding affinity and the conformational entropy of nanobody SARS-CoV-2 spike protein complexes Halina Mikolajek , Miriam Weckener , Z. Faidon Brotzakis , Jiandong Huo , Evmorfia V. Dalietou , Audrey Le Bas , Pietro Sormanni , Peter J. Harrison , Philip N. Ward , Steven Truong , Lucile Moynie , Daniel K. Clare , Maud Dumoux , Joshua Dormon , Chelsea Norman , Naveed Hussain , Vinod Vogirala , Raymond J. Owens , Michele Vendruscolo , James Naismith Proceedings Of The National Academy Of Sciences 119 DOI: 10.1073/pnas.2205412119 Diamond Proposal Number(s): [27031, 27051, 29666] Open Access Show Abstract ▼ Abstract: Camelid single-domain antibodies, also known as nanobodies, can be readily isolated from naïve libraries for specific targets but often bind too weakly to their targets to be immediately useful. Laboratory-based genetic engineering methods to enhance their affinity, termed maturation, can deliver useful reagents for different areas of biology and potentially medicine. Using the receptor binding domain (RBD) of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein and a naïve library, we generated closely related nanobodies with micromolar to nanomolar binding affinities. By analyzing the structure–activity relationship using X-ray crystallography, cryoelectron microscopy, and biophysical methods, we observed that higher conformational entropy losses in the formation of the spike protein–nanobody complex are associated with tighter binding. To investigate this, we generated structural ensembles of the different complexes from electron microscopy maps and correlated the conformational fluctuations with binding affinity. This insight guided the engineering of a nanobody with improved affinity for the spike protein.	Jul 2022
I03-Macromolecular Crystallography	Antibody escape of SARS-CoV-2 Omicron BA.4 and BA.5 from vaccine and BA.1 serum Aekkachai Tuekprakhon , Jiandong Huo , Rungtiwa Nutalai , Aiste Dijokaite-Guraliuc , Daming Zhou , Helen M. Ginn , Muneeswaran Selvaraj , Chang Liu , Alexander J. Mentzer , Piyada Supasa , Helen M. E. Duyvesteyn , Raksha Das , Donal Skelly , Thomas G. Ritter , Ali Amini , Sagida Bibi , Sandra Adele , Sile Ann Johnson , Bede Constantinides , Hermione Webster , Nigel Temperton , Paul Klenerman , Eleanor Barnes , Susanna J. Dunachie , Derrick Crook , Andrew J. Pollard , Teresa Lambe , Philip Goulder , Neil G. Paterson , Mark A. Williams , David R. Hall , Elizabeth E. Fry , Juthathip Mongkolsapaya , Jingshan Ren , David I. Stuart , Gavin R. Screaton , Christopher Conlon , Alexandra Deeks , John Frater , Lisa Frending , Siobhan Gardiner , Anni Jämsén , Katie Jeffery , Tom Malone , Eloise Phillips , Lucy Rothwell , Lizzie Stafford Cell DOI: 10.1016/j.cell.2022.06.005 Diamond Proposal Number(s): [27009] Open Access Show Abstract ▼ Abstract: The Omicron lineage of SARS-CoV-2, first described in November 2021, spread rapidly to become globally dominant and has split into a number of sub-lineages. BA.1 dominated the initial wave but has been replaced by BA.2 in many countries. Recent sequencing from South Africa’s Gauteng region uncovered two new sub-lineages, BA.4 and BA.5 which are taking over locally, driving a new wave. BA.4 and BA.5 contain identical spike sequences and, although closely related to BA.2, contain further mutations in the receptor binding domain of spike. Here, we study the neutralization of BA.4/5 using a range of vaccine and naturally immune serum and panels of monoclonal antibodies. BA.4/5 shows reduced neutralization by serum from triple AstraZeneca or Pfizer vaccinated individuals compared to BA.1 and BA.2. Furthermore, using serum from BA.1 vaccine breakthrough infections there are likewise, significant reductions in the neutralization of BA.4/5, raising the possibility of repeat Omicron infections.	Jun 2022
NONE-No attached Diamond beamline	Pathogen-sugar interactions revealed by universal saturation transfer analysis Charles J. Buchanan , Ben Gaunt , Peter J. Harrison , Yun Yang , Jiwei Liu , Aziz Khan , Andrew M. Giltrap , Audrey Le Bas , Philip N. Ward , Kapil Gupta , Maud Dumoux , Tiong Kit Tan , Lisa Schimaski , Sergio Daga , Nicola Picchiotti , Margherita Baldassarri , Elisa Benetti , Chiara Fallerini , Francesca Fava , Annarita Giliberti , Panagiotis I. Koukos , Matthew J. Davy , Abirami Lakshminarayanan , Xiaochao Xue , Georgios Papadakis , Lachlan P. Deimel , Virgínia Casablancas-Antràs , Timothy D. W. Claridge , Alexandre M. J. J. Bonvin , Quentin J. Sattentau , Simone Furini , Marco Gori , Jiandong Huo , Raymond J. Owens , Christiane Schaffitzel , Imre Berger , Alessandra Renieri , James H. Naismith , Andrew J. Baldwin , Benjamin G. Davis Science 479 DOI: 10.1126/science.abm3125 Open Access Show Abstract ▼ Abstract: Many pathogens exploit host cell-surface glycans. However, precise analyses of glycan ligands binding with heavily-modified pathogen proteins can be confounded by overlapping sugar signals and/or compound with known experimental constraints. ‘Universal saturation transfer analysis’ (uSTA) builds on existing nuclear magnetic resonance spectroscopy to provide an automated workflow for quantitating protein-ligand interactions. uSTA reveals that early-pandemic, B-origin lineage SARS-CoV-2 spike trimer binds sialoside sugars in an ‘end-on’ manner. uSTA-guided modelling and a high-resolution cryo-electron microscopy structure implicate the spike N-terminal domain (NTD) and confirm end-on binding. This finding rationalizes the effect of NTD mutations that abolish sugar-binding in SARS CoV 2 variants of concern. Together with genetic variance analyses in early pandemic patient cohorts, this binding implicates a sialylated polylactosamine motif found on tetraantennary N-linked glycoproteins in deeper human lung as potentially relevant to virulence and/or zoonosis.	Jun 2022
I03-Macromolecular Crystallography I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Production and crystallization of nanobodies in complex with the receptor binding domain of the SARS-COV-2 spike protein Audrey Le Bas , Halina Mikolajek , Jiandong Huo , Chelsea Norman , Joshua Dormon , James Naismith , Raymond Owens Bio-Protocol 12 DOI: 10.21769/BioProtoc.4406 Diamond Proposal Number(s): [27031] Show Abstract ▼ Abstract: The receptor binding domain (RBD) of the spike protein of SARS-CoV-2 binds angiotensin converting enzyme-2 (ACE-2) on the surface of epithelial cells, leading to fusion, and entry of the virus into the cell. This interaction can be blocked by the binding of llama-derived nanobodies (VHHs) to the RBD, leading to virus neutralisation. Structural analysis of VHH-RBD complexes by X-ray crystallography enables VHH epitopes to be precisely mapped, and the effect of variant mutations to be interpreted and predicted. Key to this is a protocol for the reproducible production and crystallization of the VHH-RBD complexes. Based on our experience, we describe a workflow for expressing and purifying the proteins, and the screening conditions for generating diffraction quality crystals of VHH-RBD complexes. Production and crystallization of protein complexes takes approximately twelve days, from construction of vectors to harvesting and freezing crystals for data collection.	May 2022
I03-Macromolecular Crystallography I04-Macromolecular Crystallography Krios I-Titan Krios I at Diamond	Potent cross-reactive antibodies following Omicron breakthrough in vaccinees Rungtiwa Nutalai , Daming Zhou , Aekkachai Tuekprakhon , Helen M. Ginn , Piyada Supasa , Chang Liu , Jiandong Huo , Alexander J. Mentzer , Helen M. E. Duyvesteyn , Aiste Dijokaite-Guraliuc , Donal Skelly , Thomas G. Ritter , Ali Amini , Sagida Bibi , Sandra Adele , Sile Ann Johnson , Bede Constantinides , Hermione Webster , Nigel Temperton , Paul Klenerman , Eleanor Barnes , Susanna J. Dunachie , Derrick Crook , Andrew J. Pollard , Teresa Lambe , Philip Goulder , Neil G. Paterson , Mark A. Williams , David R. Hall , Juthathip Mongkolsapaya , Elizabeth E. Fry , Wanwisa Dejnirattisai , Jingshan Ren , David I. Stuart , Gavin R. Screaton , Christopher Conlon , Alexandra Deeks , John Frater , Lisa Frending , Siobhan Gardiner , Anni Jämsén , Katie Jeffery , Tom Malone , Eloise Phillips , Lucy Rothwell , Lizzie Stafford Cell DOI: 10.1016/j.cell.2022.05.014 Diamond Proposal Number(s): [27009, 26983] Open Access Show Abstract ▼ Abstract: Highly transmissible Omicron variants of SARS-CoV-2 currently dominate globally. Here, we compare neutralization of Omicron BA.1, BA.1.1 and BA.2. BA.2 RBD has slightly higher ACE2 affinity than BA.1 and slightly reduced neutralization by vaccine serum, possibly associated with its increased transmissibility. Neutralization differences between sub-lineages for mAbs (including therapeutics) mostly arise from variation in residues bordering the ACE2 binding site, however, more distant mutations S371F (BA.2) and R346K (BA.1.1) markedly reduce neutralization by therapeutic antibody Vir-S309. In-depth structure-and-function analyses of 27 potent RBD-binding mAbs isolated from vaccinated volunteers following breakthrough Omicron-BA.1 infection reveals that they are focussed in two main clusters within the RBD, with potent right-shoulder antibodies showing increased prevalence. Selection and somatic maturation have optimized antibody potency in less-mutated epitopes and recovered potency in highly mutated epitopes. All 27 mAbs potently neutralize early pandemic strains and many show broad reactivity with variants of concern.	May 2022
I03-Macromolecular Crystallography	SARS-CoV-2 Omicron-B.1.1.529 leads to widespread escape from neutralizing antibody responses Wanwisa Dejnirattisai , Jiandong Huo , Daming Zhou , Jiří Zahradník , Piyada Supasa , Chang Liu , Helen M. E. Duyvesteyn , Helen M. Ginn , Alexander J. Mentzer , Aekkachai Tuekprakhon , Rungtiwa Nutalai , Beibei Wang , Aiste Dijokaite , Suman Khan , Ori Avinoam , Mohammad Bahar , Donal Skelly , Sandra Adele , Sile Ann Johnson , Ali Amini , Thomas Ritter , Chris Mason , Christina Dold , Daniel Pan , Sara Assadi , Adam Bellass , Nikki Omo-Dare , David Koeckerling , Amy Flaxman , Daniel Jenkin , Parvinder K. Aley , Merryn Voysey , Sue Ann Costa Clemens , Felipe Gomes Naveca , Valdinete Nascimento , Fernanda Nascimento , Cristiano Fernandes Da Costa , Paola Cristina Resende , Alex Pauvolid-Correa , Marilda M. Siqueira , Vicky Baillie , Natali Serafin , Gaurav Kwatra , Kelly Da Silva , Shabir A. Madhi , Marta C. Nunes , Tariq Malik , Peter J. M. Openshaw , J. Kenneth Baillie , Malcolm G. Semple , Alain R. Townsend , Kuan-Ying A. Huang , Tiong Kit Tan , Miles W. Carroll , Paul Klenerman , Eleanor Barnes , Susanna J. Dunachie , Bede Constantinides , Hermione Webster , Derrick Crook , Andrew J. Pollard , Teresa Lambe , Neil G. Paterson , Mark A. Williams , David R. Hall , Elizabeth E. Fry , Juthathip Mongkolsapaya , Jingshan Ren , Gideon Schreiber , David I. Stuart , Gavin R. Screaton Cell DOI: 10.1016/j.cell.2021.12.046 Diamond Proposal Number(s): [27009] Show Abstract ▼ Abstract: On the 24th November 2021 the sequence of a new SARS CoV-2 viral isolate Omicron-B.1.1.529 was announced, containing far more mutations in Spike (S) than previously reported variants. Neutralization titres of Omicron by sera from vaccinees and convalescent subjects infected with early pandemic as well as Alpha, Beta, Gamma, Delta are substantially reduced or fail to neutralize. Titres against Omicron are boosted by third vaccine doses and are high in cases both vaccinated and infected by Delta. Mutations in Omicron knock out or substantially reduce neutralization by most of a large panel of potent monoclonal antibodies and antibodies under commercial development. Omicron S has structural changes from earlier viruses, combining mutations conferring tight binding to ACE2 to unleash evolution driven by immune escape, leading to a large number of mutations in the ACE2 binding site which rebalance receptor affinity to that of early pandemic viruses.	Jan 2022
I03-Macromolecular Crystallography I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	A potent SARS-CoV-2 neutralising nanobody shows therapeutic efficacy in the Syrian golden hamster model of COVID-19 Jiandong Huo , Halina Mikolajek , Audrey Le Bas , Jordan J. Clark , Parul Sharma , Anja Kipar , Joshua Dormon , Chelsea Norman , Miriam Weckener , Daniel K. Clare , Peter J. Harrison , Julia A. Tree , Karen R. Buttigieg , Francisco J. Salguero , Robert Watson , Daniel Knott , Oliver Carnell , Didier Ngabo , Michael J. Elmore , Susan Fotheringham , Adam Harding , Lucile Moynie , Philip N. Ward , Maud Dumoux , Tessa Prince , Yper Hall , Julian A. Hiscox , Andrew Owen , William James , Miles W. Carroll , James P. Stewart , James Naismith , Raymond Owens Nature Communications 12 DOI: 10.1038/s41467-021-25480-z Diamond Proposal Number(s): [27031] Open Access Show Abstract ▼ Abstract: SARS-CoV-2 remains a global threat to human health particularly as escape mutants emerge. There is an unmet need for effective treatments against COVID-19 for which neutralizing single domain antibodies (nanobodies) have significant potential. Their small size and stability mean that nanobodies are compatible with respiratory administration. We report four nanobodies (C5, H3, C1, F2) engineered as homotrimers with pmolar affinity for the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Crystal structures show C5 and H3 overlap the ACE2 epitope, whilst C1 and F2 bind to a different epitope. Cryo Electron Microscopy shows C5 binding results in an all down arrangement of the Spike protein. C1, H3 and C5 all neutralize the Victoria strain, and the highly transmissible Alpha (B.1.1.7 first identified in Kent, UK) strain and C1 also neutralizes the Beta (B.1.35, first identified in South Africa). Administration of C5-trimer via the respiratory route showed potent therapeutic efficacy in the Syrian hamster model of COVID-19 and separately, effective prophylaxis. The molecule was similarly potent by intraperitoneal injection.	Sep 2021
	A COVID-19 vaccine candidate using SpyCatcher multimerization of the SARS-CoV-2 spike protein receptor-binding domain induces potent neutralising antibody responses Tiong Kit Tan , Pramila Rijal , Rolle Rahikainen , Anthony H. Keeble , Lisa Schimanski , Saira Hussain , Ruth Harvey , Jack W. P. Hayes , Jane C. Edwards , Rebecca K. Mclean , Veronica Martini , Miriam Pedrera , Nazia Thakur , Carina Conceicao , Isabelle Dietrich , Holly Shelton , Anna Ludi , Ginette Wilsden , Clare Browning , Adrian K. Zagrajek , Dagmara Bialy , Sushant Bhat , Phoebe Stevenson-Leggett , Philippa Hollinghurst , Matthew Tully , Katy Moffat , Chris Chiu , Ryan Waters , Ashley Gray , Mehreen Azhar , Valerie Mioulet , Joseph Newman , Amin S. Asfor , Alison Burman , Sylvia Crossley , John A. Hammond , Elma Tchilian , Bryan Charleston , Dalan Bailey , Tobias J. Tuthill , Simon P. Graham , Helen M. E. Duyvesteyn , Tomas Malinauskas , Jiandong Huo , Julia A. Tree , Karen R. Buttigieg , Raymond J. Owens , Miles W. Carroll , Rodney S. Daniels , John W. Mccauley , David I. Stuart , Kuan-Ying A. Huang , Mark Howarth , Alain R. Townsend Nature Communications 12 DOI: 10.1038/s41467-020-20654-7 Open Access Show Abstract ▼ Abstract: There is need for effective and affordable vaccines against SARS-CoV-2 to tackle the ongoing pandemic. In this study, we describe a protein nanoparticle vaccine against SARS-CoV-2. The vaccine is based on the display of coronavirus spike glycoprotein receptor-binding domain (RBD) on a synthetic virus-like particle (VLP) platform, SpyCatcher003-mi3, using SpyTag/SpyCatcher technology. Low doses of RBD-SpyVLP in a prime-boost regimen induce a strong neutralising antibody response in mice and pigs that is superior to convalescent human sera. We evaluate antibody quality using ACE2 blocking and neutralisation of cell infection by pseudovirus or wild-type SARS-CoV-2. Using competition assays with a monoclonal antibody panel, we show that RBD-SpyVLP induces a polyclonal antibody response that recognises key epitopes on the RBD, reducing the likelihood of selecting neutralisation-escape mutants. Moreover, RBD-SpyVLP is thermostable and can be lyophilised without losing immunogenicity, to facilitate global distribution and reduce cold-chain dependence. The data suggests that RBD-SpyVLP provides strong potential to address clinical and logistic challenges of the COVID-19 pandemic.	Jan 2021

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