NONE-No attached Diamond beamline
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H. T. Henry
Chan
,
Marc A.
Moesser
,
Rebecca K.
Walters
,
Tika R.
Malla
,
Rebecca M.
Twidale
,
Tobias
John
,
Helen M.
Deeks
,
Tristan
Johnston-Wood
,
Victor
Mikhailov
,
Richard B.
Sessions
,
William
Dawson
,
Eidarus
Salah
,
Petra
Lukacik
,
Claire
Strain-Damerell
,
C. David
Owen
,
Takahito
Nakajima
,
Katarzyna
Świderek
,
Alessio
Lodola
,
Vicent
Moliner
,
David R.
Glowacki
,
James
Spencer
,
Martin A.
Walsh
,
Christopher J.
Schofield
,
Luigi
Genovese
,
Deborah K.
Shoemark
,
Adrian J.
Mulholland
,
Fernanda
Duarte
,
Garrett M.
Morris
Open Access
Abstract: The main protease (Mpro) of SARS-CoV-2 is central to viral maturation and is a promising drug target, but little is known about structural aspects of how it binds to its 11 natural cleavage sites. We used biophysical and crystallographic data and an array of biomolecular simulation techniques, including automated docking, molecular dynamics (MD) and interactive MD in virtual reality, QM/MM, and linear-scaling DFT, to investigate the molecular features underlying recognition of the natural Mpro substrates. We extensively analysed the subsite interactions of modelled 11-residue cleavage site peptides, crystallographic ligands, and docked COVID Moonshot-designed covalent inhibitors. Our modelling studies reveal remarkable consistency in the hydrogen bonding patterns of the natural Mpro substrates, particularly on the N-terminal side of the scissile bond. They highlight the critical role of interactions beyond the immediate active site in recognition and catalysis, in particular plasticity at the S2 site. Building on our initial Mpro-substrate models, we used predictive saturation variation scanning (PreSaVS) to design peptides with improved affinity. Non-denaturing mass spectrometry and other biophysical analyses confirm these new and effective ‘peptibitors’ inhibit Mpro competitively. Our combined results provide new insights and highlight opportunities for the development of Mpro inhibitors as anti-COVID-19 drugs.
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Oct 2021
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I04-1-Macromolecular Crystallography (fixed wavelength)
|
Muhamamd
Faheem
,
Napoleao
Fonseca Valadares
,
Jose
Brandao-Neto
,
Domenico
Bellini
,
Patrick
Collins
,
Nicholas M.
Pearce
,
Louise
Bird
,
Juliana
Torini De Souza
,
Raymond
Owens
,
Humberto
Pereira
,
Frank
Von Delft
,
João Alexandre Ribeiro Gonçalves
Barbosa
Diamond Proposal Number(s):
[11175]
Open Access
Abstract: Several Schistosoma species cause Schistosomiasis, an endemic disease in 78 countries that is ranked second amongst the parasitic diseases in terms of its socioeconomic impact and human health importance. The drug recommended for treatment by the WHO is praziquantel (PZQ), but there are concerns associated with PZQ, such as the lack of information about its exact mechanism of action, its high price, its effectiveness – which is limited to the parasite’s adult form – and reports of resistance. The parasites lack the de novo purine pathway, rendering them dependent on the purine salvage pathway or host purine bases for nucleotide synthesis. Thus, the Schistosoma purine salvage pathway is an attractive target for the development of necessary and selective new drugs. In this study, the purine nucleotide phosphorylase II (PNP2), a new isoform of PNP1, was submitted to a high-throughput fragment-based hit discovery using a crystallographic screening strategy. PNP2 was crystallized and crystals were soaked with 827 fragments, a subset of the Maybridge 1000 library. X-ray diffraction data was collected and structures were solved. Out of 827-screened fragments we have obtained a total of 19 fragments that show binding to PNP2. 14 of these fragments bind to the active site of PNP2, while five were observed in three other sites. Here we present the first fragment screening against PNP2.
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Sep 2021
|
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I04-1-Macromolecular Crystallography (fixed wavelength)
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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
Diamond Proposal Number(s):
[26998]
Open Access
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.
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Aug 2021
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I04-1-Macromolecular Crystallography (fixed wavelength)
|
Open Access
Abstract: Plasmodium falciparum invades erythrocytes and extensively modifies them in a manner that increases the virulence of this malaria parasite. A single heat-shock 70 kDa-type chaperone, PfHsp70-x, is among the parasite proteins exported to the host cell. PfHsp70-x assists in the formation of a key protein complex that underpins parasite virulence and supports parasite growth during febrile episodes. Previous work resolved the crystallographic structures of the PfHsp70-x ATPase and substrate-binding domains, and showed them to be highly similar to those of their human counterparts. Here, 233 chemical fragments were screened for binding to the PfHsp70-x ATPase domain, resulting in three crystallographic structures of this domain in complex with ligands. Two binding sites were identified, with most ligands binding proximal to the ATPase nucleotide-binding pocket. Although amino acids participating in direct ligand interactions are conserved between the parasite and human erythrocytic chaperones, one nonconserved residue is also present near the ligand. This work suggests that PfHsp70-x features binding sites that may be exploitable by small-molecule ligands towards the specific inhibition of the parasite chaperone.
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Aug 2021
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Hector
Newman
,
Alen
Krajnc
,
Domenico
Bellini
,
Charles J.
Eyermann
,
Grant A.
Boyle
,
Neil
Paterson
,
Katherine E.
Mcauley
,
Robert
Lesniak
,
Mukesh
Gangar
,
Frank
Von Delft
,
Jurgen
Brem
,
Kelly
Chibale
,
Christopher J.
Schofield
,
Christopher G.
Dowson
Diamond Proposal Number(s):
[17884]
Open Access
Abstract: The effectiveness of β-lactam antibiotics is increasingly compromised by β-lactamases. Boron-containing inhibitors are potent serine-β-lactamase inhibitors, but the interactions of boron-based compounds with the penicillin-binding protein (PBP) β-lactam targets have not been extensively studied. We used high-throughput X-ray crystallography to explore reactions of a boron-containing fragment set with the Pseudomonas aeruginosa PBP3 (PaPBP3). Multiple crystal structures reveal that boronic acids react with PBPs to give tricovalently linked complexes bonded to Ser294, Ser349, and Lys484 of PaPBP3; benzoxaboroles react with PaPBP3 via reaction with two nucleophilic serines (Ser294 and Ser349) to give dicovalently linked complexes; and vaborbactam reacts to give a monocovalently linked complex. Modifications of the benzoxaborole scaffold resulted in a moderately potent inhibition of PaPBP3, though no antibacterial activity was observed. Overall, the results further evidence the potential for the development of new classes of boron-based antibiotics, which are not compromised by β-lactamase-driven resistance.
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Jul 2021
|
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I04-1-Macromolecular Crystallography (fixed wavelength)
|
Diamond Proposal Number(s):
[17334]
Open Access
Abstract: Tubulin plays essential roles in vital cellular activities and is the target of a wide range of proteins and ligands. Here, using a combined computational and crystallographic fragment screening approach, we addressed the question of how many binding sites exist in tubulin. We identified 27 distinct sites, of which 11 have not been described previously, and analyzed their relationship to known tubulin–protein and tubulin–ligand interactions. We further observed an intricate pocket communication network and identified 56 chemically diverse fragments that bound to 10 distinct tubulin sites. Our results offer a unique structural basis for the development of novel small molecules for use as tubulin modulators in basic research applications or as drugs. Furthermore, our method lays down a framework that may help to discover new pockets in other pharmaceutically important targets and characterize them in terms of chemical tractability and allosteric modulation.
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May 2021
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I04-1-Macromolecular Crystallography (fixed wavelength)
|
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
Diamond Proposal Number(s):
[27001, 18145, 27963]
Open Access
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.
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May 2021
|
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I04-1-Macromolecular Crystallography (fixed wavelength)
|
José T.
Moreira-Filho
,
Arthur C.
Silva
,
Rafael F.
Dantas
,
Barbara F.
Gomes
,
Lauro R.
De Souza Neto
,
Jose
Brandao-Neto
,
Raymond J.
Owens
,
Nicholas
Furnham
,
Bruno J.
Neves
,
Floriano P.
Silva-Junior
,
Carolina H.
Andrade
Diamond Proposal Number(s):
[16978]
Open Access
Abstract: Schistosomiasis is a parasitic disease caused by trematode worms of the genus Schistosoma and affects over 200 million people worldwide. The control and treatment of this neglected tropical disease is based on a single drug, praziquantel, which raises concerns about the development of drug resistance. This, and the lack of efficacy of praziquantel against juvenile worms, highlights the urgency for new antischistosomal therapies. In this review we focus on innovative approaches to the identification of antischistosomal drug candidates, including the use of automated assays, fragment-based screening, computer-aided and artificial intelligence-based computational methods. We highlight the current developments that may contribute to optimizing research outputs and lead to more effective drugs for this highly prevalent disease, in a more cost-effective drug discovery endeavor.
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May 2021
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NONE-No attached Diamond beamline
|
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
Open Access
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.
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May 2021
|
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I04-1-Macromolecular Crystallography (fixed wavelength)
|
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
Diamond Proposal Number(s):
[27001]
Open Access
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.
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Apr 2021
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