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

Instruments / Technical Area	Publication Details	Published
I04-1-Macromolecular Crystallography (fixed wavelength)	Structure, mechanism and crystallographic fragment screening of the SARS-CoV-2 NSP13 helicase Joseph A. Newman , Alice Douangamath , Setayesh Yadzani , Yuliana Yosaatmadja , Anthony Aimon , Jose Brandao-Neto , Louise Dunnett , Tyler Gorrie-Stone , Rachael Skyner , Daren Fearon , Matthieu Schapira , Frank Von Delft , Opher Gileadi Nature Communications 12 DOI: 10.1038/s41467-021-25166-6 Diamond Proposal Number(s): [26998] Open Access Show Abstract ▼ Abstract: There is currently a lack of effective drugs to treat people infected with SARS-CoV-2, the cause of the global COVID-19 pandemic. The SARS-CoV-2 Non-structural protein 13 (NSP13) has been identified as a target for anti-virals due to its high sequence conservation and essential role in viral replication. Structural analysis reveals two “druggable” pockets on NSP13 that are among the most conserved sites in the entire SARS-CoV-2 proteome. Here we present crystal structures of SARS-CoV-2 NSP13 solved in the APO form and in the presence of both phosphate and a non-hydrolysable ATP analog. Comparisons of these structures reveal details of conformational changes that provide insights into the helicase mechanism and possible modes of inhibition. To identify starting points for drug development we have performed a crystallographic fragment screen against NSP13. The screen reveals 65 fragment hits across 52 datasets opening the way to structure guided development of novel antiviral agents.	Aug 2021
I04-1-Macromolecular Crystallography (fixed wavelength)	A crystallographic snapshot of SARS-CoV-2 main protease maturation process G. D. Noske , A. M. Nakamura , V. O. Gawriljuk , R. S. Fernandes , G. M. A. Lima , H. V. D. Rosa , H. D. Pereira , A. C. M. Zeri , A. A. F. Z. Nascimento , M. C. L. C. Freire , D. Fearon , A. Douangamath , F. Von Delft , G. Oliva , A. S. Godoy Journal Of Molecular Biology DOI: 10.1016/j.jmb.2021.167118 Diamond Proposal Number(s): [27963] Show Abstract ▼ Abstract: SARS-CoV-2 is the causative agent of COVID-19. The dimeric form of the viral Mpro is responsible for the cleavage of the viral polyprotein in 11 sites, including its own N and C-terminus. The lack of structural information for intermediary forms of Mpro is a setback for the understanding its self-maturation process. Herein, we used X-ray crystallography combined with biochemical data to characterize multiple forms of SARS-CoV-2 Mpro. For the immature form, we show that extra N-terminal residues caused conformational changes in the positioning of domain-three over the active site, hampering the dimerization and diminishing its activity. We propose that this form preludes the cis and trans-cleavage of N-terminal residues. Using fragment screening, we probe new cavities in this form which can be used to guide therapeutic development. Furthermore, we characterized a serine site-directed mutant of the Mpro bound to its endogenous N and C-terminal residues during dimeric association stage of the maturation process. We suggest this form is a transitional state during the C-terminal trans-cleavage. This data sheds light in the structural modifications of the SARS-CoV-2 main protease during its self-maturation process.	Jun 2021
I04-1-Macromolecular Crystallography (fixed wavelength)	An automatic pipeline for the design of irreversible derivatives identifies a potent SARS-CoV-2 Mpro inhibitor Daniel Zaidman , Paul Gehrtz , Mihajlo Filep , Daren Fearon , Ronen Gabizon , Alice Douangamath , Jaime Prilusky , Shirly Duberstein , Galit Cohen , C. David Owen , Efrat Resnick , Claire Strain-Damerell , Petra Lukacik , Haim Barr , Martin A. Walsh , Frank Von Delft , Nir London Cell Chemical Biology 63 DOI: 10.1016/j.chembiol.2021.05.018 Diamond Proposal Number(s): [18145, 27963] Show Abstract ▼ Abstract: Designing covalent inhibitors is increasingly important, although it remains challenging. Here, we present covalentizer, a computational pipeline for identifying irreversible inhibitors based on structures of targets with non-covalent binders. Through covalent docking of tailored focused libraries, we identify candidates that can bind covalently to a nearby cysteine while preserving the interactions of the original molecule. We found ∼11,000 cysteines proximal to a ligand across 8,386 complexes in the PDB. Of these, the protocol identified 1,553 structures with covalent predictions. In a prospective evaluation, five out of nine predicted covalent kinase inhibitors showed half-maximal inhibitory concentration (IC50) values between 155 nM and 4.5 μM. Application against an existing SARS-CoV Mpro reversible inhibitor led to an acrylamide inhibitor series with low micromolar IC50 values against SARS-CoV-2 Mpro. The docking was validated by 12 co-crystal structures. Together these examples hint at the vast number of covalent inhibitors accessible through our protocol.	Jun 2021
I04-1-Macromolecular Crystallography (fixed wavelength) I24-Microfocus Macromolecular Crystallography	Bispecific repurposed medicines targeting the viral and immunological arms of COVID-19 Martin A. Redhead , C. David Owen , Lennart Brewitz , Amelia H. Collette , Petra Lukacik , Claire Strain-Damerell , Sean W. Robinson , Patrick M. Collins , Philipp Schäfer , Mark Swindells , Chris J. Radoux , Iva Navratilova Hopkins , Daren Fearon , Alice Douangamath , Frank Von Delft , Tika R. Malla , Laura Vangeel , Thomas Vercruysse , Jan Thibaut , Pieter Leyssen , Tu-Trinh Nguyen , Mitchell Hull , Anthony Tumber , David J. Hallett , Christopher J. Schofield , David I. Stuart , Andrew L. Hopkins , Martin A. Walsh Scientific Reports 11 DOI: 10.1038/s41598-021-92416-4 Open Access Show Abstract ▼ Abstract: Effective agents to treat coronavirus infection are urgently required, not only to treat COVID-19, but to prepare for future outbreaks. Repurposed anti-virals such as remdesivir and human anti-inflammatories such as barcitinib have received emergency approval but their overall benefits remain unclear. Vaccines are the most promising prospect for COVID-19, but will need to be redeveloped for any future coronavirus outbreak. Protecting against future outbreaks requires the identification of targets that are conserved between coronavirus strains and amenable to drug discovery. Two such targets are the main protease (Mpro) and the papain-like protease (PLpro) which are essential for the coronavirus replication cycle. We describe the discovery of two non-antiviral therapeutic agents, the caspase-1 inhibitor SDZ 224015 and Tarloxotinib that target Mpro and PLpro, respectively. These were identified through extensive experimental screens of the drug repurposing ReFRAME library of 12,000 therapeutic agents. The caspase-1 inhibitor SDZ 224015, was found to be a potent irreversible inhibitor of Mpro (IC50 30 nM) while Tarloxotinib, a clinical stage epidermal growth factor receptor inhibitor, is a sub micromolar inhibitor of PLpro (IC50 300 nM, Ki 200 nM) and is the first reported PLpro inhibitor with drug-like properties. SDZ 224015 and Tarloxotinib have both undergone safety evaluation in humans and hence are candidates for COVID-19 clinical evaluation.	Jun 2021
I04-1-Macromolecular Crystallography (fixed wavelength)	Exploring protein hotspots by optimized fragment pharmacophores Dávid Bajusz , Warren S. Wade , Grzegorz Satała , Andrzej J. Bojarski , Janez Ilaš , Jessica Ebner , Florian Grebien , Henrietta Papp , Ferenc Jakab , Alice Douangamath , Daren Fearon , Frank Von Delft , Marion Schuller , Ivan Ahel , Amanda Wakefield , Sándor Vajda , János Gerencsér , Péter Pallai , György M. Keserű Nature Communications 12 DOI: 10.1038/s41467-021-23443-y Diamond Proposal Number(s): [27001, 18145, 27963] Open Access Show Abstract ▼ Abstract: Fragment-based drug design has introduced a bottom-up process for drug development, with improved sampling of chemical space and increased effectiveness in early drug discovery. Here, we combine the use of pharmacophores, the most general concept of representing drug-target interactions with the theory of protein hotspots, to develop a design protocol for fragment libraries. The SpotXplorer approach compiles small fragment libraries that maximize the coverage of experimentally confirmed binding pharmacophores at the most preferred hotspots. The efficiency of this approach is demonstrated with a pilot library of 96 fragment-sized compounds (SpotXplorer0) that is validated on popular target classes and emerging drug targets. Biochemical screening against a set of GPCRs and proteases retrieves compounds containing an average of 70% of known pharmacophores for these targets. More importantly, SpotXplorer0 screening identifies confirmed hits against recently established challenging targets such as the histone methyltransferase SETD2, the main protease (3CLPro) and the NSP3 macrodomain of SARS-CoV-2.	May 2021
NONE-No attached Diamond beamline	Achieving efficient fragment screening at XChem facility at Diamond Light Source Alice Douangamath , Alisa Powell , Daren Fearon , Patrick M. Collins , Romain Talon , Tobias Krojer , Rachael Skyner , Jose Brandao-Neto , Louise Dunnett , Alexandre Dias , Anthony Aimon , Nicholas M. Pearce , Conor Wild , Tyler J. Gorrie-Stone , Frank Von Delft Journal Of Visualized Experiments DOI: 10.3791/62414 Open Access Show Abstract ▼ Abstract: In fragment-based drug discovery, hundreds or often thousands of compounds smaller than ~300 Da are tested against the protein of interest to identify chemical entities that can be developed into potent drug candidates. Since the compounds are small, interactions are weak, and the screening method must therefore be highly sensitive; moreover, structural information tends to be crucial for elaborating these hits into lead-like compounds. Therefore, protein crystallography has always been a gold-standard technique, yet historically too challenging to find widespread use as a primary screen. Initial XChem experiments were demonstrated in 2014 and then trialed with academic and industrial collaborators to validate the process. Since then, a large research effort and significant beamtime have streamlined sample preparation, developed a fragment library with rapid follow-up possibilities, automated and improved the capability of I04-1 beamline for unattended data collection, and implemented new tools for data management, analysis and hit identification. XChem is now a facility for large-scale crystallographic fragment screening, supporting the entire crystals-to-deposition process, and accessible to academic and industrial users worldwide. The peer-reviewed academic user program has been actively developed since 2016, to accommodate projects from as broad a scientific scope as possible, including well-validated as well as exploratory projects. Academic access is allocated through biannual calls for peer-reviewed proposals, and proprietary work is arranged by Diamond's Industrial Liaison group. This workflow has already been routinely applied to over a hundred targets from diverse therapeutic areas, and effectively identifies weak binders (1%-30% hit rate), which both serve as high-quality starting points for compound design and provide extensive structural information on binding sites. The resilience of the process was demonstrated by continued screening of SARS-CoV-2 targets during the COVID-19 pandemic, including a 3-week turn-around for the main protease.	May 2021
I04-1-Macromolecular Crystallography (fixed wavelength)	Fragment binding to the Nsp3 macrodomain of SARS-CoV-2 identified through crystallographic screening and computational docking Marion Schuller , Galen J. Correy , Stefan Gahbauer , Daren Fearon , Taiasean Wu , Roberto Efraín Díaz , Iris D. Young , Luan Carvalho Martins , Dominique H. Smith , Ursula Schulze-Gahmen , Tristan W. Owens , Ishan Deshpande , Gregory E. Merz , Aye C. Thwin , Justin T. Biel , Jessica K. Peters , Michelle Moritz , Nadia Herrera , Huong T. Kratochvil , Anthony Aimon , James Bennett , Jose Brandao Neto , Aina E. Cohen , Alexandre Dias , Alice Douangamath , Louise Dunnett , Oleg Fedorov , Matteo P. Ferla , Martin R. Fuchs , Tyler J. Gorrie-Stone , James M. Holton , Michael G. Johnson , Tobias Krojer , George Meigs , Alisa J. Powell , Johannes Gregor Matthias Rack , Victor Rangel , Silvia Russi , Rachael E. Skyner , Clyde A. Smith , Alexei S. Soares , Jennifer L. Wierman , Kang Zhu , Peter O’brien , Natalia Jura , Alan Ashworth , John J. Irwin , Michael C. Thompson , Jason E. Gestwicki , Frank Von Delft , Brian K. Shoichet , James S. Fraser , Ivan Ahel Science Advances 7 DOI: 10.1126/sciadv.abf8711 Diamond Proposal Number(s): [27001] Open Access Show Abstract ▼ Abstract: The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) macrodomain within the nonstructural protein 3 counteracts host-mediated antiviral adenosine diphosphate–ribosylation signaling. This enzyme is a promising antiviral target because catalytic mutations render viruses nonpathogenic. Here, we report a massive crystallographic screening and computational docking effort, identifying new chemical matter primarily targeting the active site of the macrodomain. Crystallographic screening of 2533 diverse fragments resulted in 214 unique macrodomain-binders. An additional 60 molecules were selected from docking more than 20 million fragments, of which 20 were crystallographically confirmed. X-ray data collection to ultra-high resolution and at physiological temperature enabled assessment of the conformational heterogeneity around the active site. Several fragment hits were confirmed by solution binding using three biophysical techniques (differential scanning fluorimetry, homogeneous time-resolved fluorescence, and isothermal titration calorimetry). The 234 fragment structures explore a wide range of chemotypes and provide starting points for development of potent SARS-CoV-2 macrodomain inhibitors.	Apr 2021
I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography	Crystallographic and electrophilic fragment screening of the SARS-CoV-2 main protease Alice Douangamath , Daren Fearon , Paul Gehrtz , Tobias Krojer , Petra Lukacik , C. David Owen , Efrat Resnick , Claire Strain-Damerell , Anthony Aimon , Péter Ábrányi-Balogh , Jose Brandao-Neto , Anna Carbery , Gemma Davison , Alexandre Dias , Thomas D. Downes , Louise Dunnett , Michael Fairhead , James D. Firth , S. Paul Jones , Aaron Keeley , György M. Keserü , Hanna F. Klein , Mathew P. Martin , Martin M. Noble , Peter O’brien , Ailsa Powell , Rambabu N. Reddi , Rachael Skyner , Matthew Snee , Michael J. Waring , Conor Wild , Nir London , Frank Von Delft , Martin A. Walsh Nature Communications 11 DOI: 10.1038/s41467-020-18709-w Open Access Show Abstract ▼ Abstract: COVID-19, caused by SARS-CoV-2, lacks effective therapeutics. Additionally, no antiviral drugs or vaccines were developed against the closely related coronavirus, SARS-CoV-1 or MERS-CoV, despite previous zoonotic outbreaks. To identify starting points for such therapeutics, we performed a large-scale screen of electrophile and non-covalent fragments through a combined mass spectrometry and X-ray approach against the SARS-CoV-2 main protease, one of two cysteine viral proteases essential for viral replication. Our crystallographic screen identified 71 hits that span the entire active site, as well as 3 hits at the dimer interface. These structures reveal routes to rapidly develop more potent inhibitors through merging of covalent and non-covalent fragment hits; one series of low-reactivity, tractable covalent fragments were progressed to discover improved binders. These combined hits offer unprecedented structural and reactivity information for on-going structure-based drug design against SARS-CoV-2 main protease.	Oct 2020
I04-1-Macromolecular Crystallography (fixed wavelength)	In crystallo-screening for discovery of human norovirus 3C-like protease inhibitors Jingxu Guo , Alice Douangamath , Weixiao Song , Alun Coker , A. W. Edith Chan , Steve P. Wood , Jonathan B. Cooper , Efrat Resnick , Nir London , Frank Von Delft Journal Of Structural Biology: X DOI: 10.1016/j.yjsbx.2020.100031 Open Access Show Abstract ▼ Abstract: Outbreaks of human epidemic nonbacterial gastroenteritis are mainly caused by noroviruses. Viral replication requires a 3C-like cysteine protease (3CLpro) which processes the 200 kDa viral polyprotein into six functional proteins. The 3CLpro has attracted much interest due to its potential as a target for antiviral drugs. A system for growing high-quality crystals of native Southampton norovirus 3CLpro (SV3CP) has been established, allowing the ligand-free crystal structure to be determined to 1.3 Å in a tetrameric state. This also allowed crystal-based fragment screening to be performed with various compound libraries, ultimately to guide drug discovery for SV3CP. A total of 19 fragments were found to bind to the protease out of the 844 which were screened. Two of the hits were located at the active site of SV3CP and showed good inhibitory activity in kinetic assays. Another 5 were found at the enzyme’s putative RNA-binding site and a further 10 were located in the symmetric central cavity of the tetramer.	Jul 2020
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	Crystal structure of the catalytic D2 domain of the AAA + ATPase p97 reveals a putative helical split‐washer‐type mechanism for substrate unfolding Lasse Stach , R. Marc Morgan , Linda Makhlouf , Alice Douangamath , Frank Von Delft , Xiaodong Zhang , Paul S. Freemont Febs Letters DOI: 10.1002/1873-3468.13667 Diamond Proposal Number(s): [16980] Show Abstract ▼ Abstract: Several pathologies have been associated with the AAA + ATPase p97, an enzyme essential to protein homeostasis. Heterozygous polymorphisms in p97 have been shown to cause neurological disease, while elevated proteotoxic stress in tumours has made p97 an attractive cancer chemotherapy target. The cellular processes reliant on p97 are well described. High‐resolution structural models of its catalytic D2 domain, however, have proved elusive, as has the mechanism by which p97 converts the energy from ATP hydrolysis into mechanical force to unfold protein substrates. Here, we describe the high‐resolution structure of the p97 D2 ATPase domain. This crystal system constitutes a valuable tool for p97 inhibitor development and identifies a potentially druggable pocket in the D2 domain. In addition, its P61 symmetry suggests a mechanism for substrate unfolding by p97.	Nov 2019

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