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60 items found (0 items not approved), displaying 1 to 10.[First/Prev] 1, 2, 3, 4, 5, 6 [Next/Last]

Instruments / Technical Area	Publication Details	Published
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength) I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Structure-guided fragment-based drug discovery at the synchrotron: screening binding sites and correlations with hotspot mapping Sherine E. Thomas , Patrick Collins , Rory Hennell James , Vitor Mendes , Sitthivut Charoensutthivarakul , Chris Radoux , Chris Abell , Anthony G. Coyne , R. Andres Floto , Frank Von Delft , Tom Blundell Philosophical Transactions Of The Royal Society A: Mathematical, Physical And Engineering Sciences 377 DOI: 10.1098/rsta.2018.0422 Open Access Show Abstract ▼ Abstract: Structure-guided drug discovery emerged in the 1970s and 1980s, stimulated by the three-dimensional structures of protein targets that became available, mainly through X-ray crystal structure analysis, assisted by the development of synchrotron radiation sources. Structures of known drugs or inhibitors were used to guide the development of leads. The growth of high-throughput screening during the late 1980s and the early 1990s in the pharmaceutical industry of chemical libraries of hundreds of thousands of compounds of molecular weight of approximately 500 Da was impressive but still explored only a tiny fraction of the chemical space of the predicted 1040 drug-like compounds. The use of fragments with molecular weights less than 300 Da in drug discovery not only decreased the chemical space needing exploration but also increased promiscuity in binding targets. Here we discuss advances in X-ray fragment screening and the challenge of identifying sites where fragments not only bind but can be chemically elaborated while retaining their positions and binding modes. We first describe the analysis of fragment binding using conventional X-ray difference Fourier techniques, with Mycobacterium abscessus SAICAR synthetase (PurC) as an example. We observe that all fragments occupy positions predicted by computational hotspot mapping. We compare this with fragment screening at Diamond Synchrotron Light Source XChem facility using PanDDA software, which identifies many more fragment hits, only some of which bind to the predicted hotspots. Many low occupancy sites identified may not support elaboration to give adequate ligand affinity, although they will likely be useful in drug discovery as ‘warm spots’ for guiding elaboration of fragments bound at hotspots. We discuss implications of these observations for fragment screening at the synchrotron sources.	Jun 2019
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	Rapid covalent-probe discovery by electrophile-fragment screening Efrat Resnick , Anthony Bradley , Jinrui Gan , Alice Douangamath , Tobias Krojer , Ritika Sethi , Paul P. Geurink , Anthony Aimon , Gabriel Amitai , Dom Bellini , James Bennett , Michael Fairhead , Oleg Fedorov , Ronen Gabizon , Jin Gan , Jingxu Guo , Alexander Plotnikov , Nava Reznik , Gian Filippo Ruda , Laura Diaz-saez , Verena M. Straub , Tamas Szommer , Srikannathasan Velupillai , Daniel Zaidman , Yanling Zhang , Alun R. Coker , Christopher G. Dowson , Haim Barr , Chu Wang , Kilian V. M. Huber , Paul E. Brennan , Huib Ovaa , Frank Von Delft , Nir London Journal Of The American Chemical Society DOI: 10.1021/jacs.9b02822 Show Abstract ▼ Abstract: Covalent probes can display unmatched potency, selectivity and duration of action; however, their discovery is challenging. In principle, fragments that can irreversibly bind their target can overcome the low affinity that limits reversible fragment screening, but such electrophilic fragments were considered non-selective and were rarely screened. We hypothesized that mild electrophiles might overcome the selectivity challenge and constructed a library of 993 mildly electrophilic fragments. We characterized this library by a new high-throughput thiol-reactivity assay and screened them against ten cysteine-containing proteins. Highly reactive and promiscuous fragments were rare and could be easily eliminated. By contrast, we found hits for most targets. Combining our approach with high-throughput crystallography allowed rapid progression to potent and selective probes for two enzymes, the deubiquitinase OTUB2 and the pyrophosphatase NUDT7. No inhibitors were previously known for either. This study highlights the potential of electrophile-fragment screening as a practical and efficient tool for covalent-ligand discovery.	May 2019
I04-1-Macromolecular Crystallography (fixed wavelength)	Construction of a shape‐diverse fragment set: Design, synthesis and screen against Aurora‐A kinase Rong Zhang , Patrick J. Mcintyre , Patrick M. Collins , Daniel J. Foley , Christopher Arthur , Frank Von Delft , Richard Bayliss , Stuart Warriner , Adam Nelson Chemistry – A European Journal 73 DOI: 10.1002/chem.201900815 Diamond Proposal Number(s): [14331] Show Abstract ▼ Abstract: Historically, chemists have explored chemical space in a highly uneven and unsystematic manner. As an example, the shape diversity of existing fragment sets does not generally reflect that of all theoretically possible fragments. To assess experimentally the added value of increased three dimensionality, a shape‐diverse fragment set was designed and collated. The set was assembled by both using commercially available fragments and harnessing unified synthetic approaches to sp3‐rich molecular scaffolds. The resulting set of 80 fragments was highly three‐dimensional, and its shape diversity was significantly enriched by twenty synthesised fragments. The fragment set was screened by high‐throughput protein crystallography against Aurora‐A kinase, revealing four hits that targeted the binding site of allosteric regulators. In the longer term, it is envisaged that the fragment set could be screened against a range of functionally diverse proteins, allowing the added value of more shape‐diverse screening collections to be more fully assessed.	Apr 2019
	Role of protein structure in variant annotation: structural insight of mutations causing 6-pyruvoyl-tetrahydropterin synthase deficiency Joao R. C. Muniz , Natalie Wing-sum Szeto , Rebecca Frise , Wen Hwa Lee , Xian-song Wang , Beat Thöny , Nastassja Himmelreich , Nenad Blau , Kwang-jen Hsiao , Tze-tze Liu , Opher Gileadi , Udo Oppermann , Frank Von Delft , Wyatt Yue , Nelson Leung-sang Tang Pathology DOI: 10.1016/j.pathol.2018.11.011 Show Abstract ▼ Abstract: Genetic defects on 6-pyruvoyl-tetrahydropterin synthase (PTPS) are the most prevalent cause of hyperphenylalaninaemia not due to phenylalanine hydrolyase deficiency (phenylketonuria). PTPS catalyses the second step of tetrahydrobiopterin (BH4) cofactor biosynthesis, and its deficiency represents the most common form of BH4 deficiency. Untreated PTPS deficiency results in depletion of the neurotransmitters dopamine, catecholamine and serotonin causing neurological symptoms. We archived reported missense variants of the PTS gene. Common in silico algorithms were used to predict the effects of such variants, and substantial proportions (up to 19%) of the variants were falsely classified as benign or uncertain. We have determined the crystal structure of the human PTPS hexamer, allowing another level of interpretation to understand the potential deleterious consequences of the variants from a structural perspective. The in silico and structure approaches appear to be complimentary and may provide new insights that are not available from each alone. Information from the protein structure suggested that the variants affecting amino acid residues required for interaction between monomeric subunits of the PTPS hexamer were those misclassified as benign by in silico algorithms. Our findings illustrate the important utility of 3D protein structure in interpretation of variants and also current limitations of in silico prediction algorithms. However, software to analyse mutation in the perspective of 3D protein structure is far less readily available than other in silico prediction tools.	Mar 2019
	Targeting the small GTPase superfamily through their regulatory proteins Janine L. Gray , Frank Von Delft , Paul Brennan Angewandte Chemie International Edition DOI: 10.1002/anie.201900585 Show Abstract ▼ Abstract: The Ras superfamily of small GTPases are guanine nucleotide dependent switches essential for numerous cellular processes. Mutations or dysregulation of these proteins are associated with many diseases, but unsuccessful attempts to target the small GTPases directly have resulted in them being classed as ‘undruggable’. The GTP dependent signaling of these proteins is controlled by their regulators; guanine nucleotide exchange factors (GEFs), GTPase activating proteins (GAPs), and in the Rho and Rab subfamilies, guanine nucleotide dissociation inhibitors (GDIs). This review covers the recent small molecule and biologics strategies to target the small GTPases through their regulators. It seeks to critically re‐evaluate recent chemical biology practice, such as the presence of PAINs motifs and the cell‐based readout using compounds that are weakly potent or of unknown specificity. It highlights the vast scope of potential approaches for targeting the small GTPases in the future through their regulatory proteins.	Mar 2019
	Achieving a good crystal system for crystallographic x-ray fragment screening Patrick M. Collins , Alice Douangamath , Romain Talon , Alexandre Dias , Jose Brandao-neto , Tobias Krojer , Frank Von Delft DOI: 10.1016/bs.mie.2018.09.027 Show Abstract ▼ Abstract: The XChem facility at Diamond Light Source offers fragment screening by X-ray crystallography as a general access user program. The main advantage of X-ray crystallography as a primary fragment screen is that it yields directly the location and pose of the fragment hits, whether within pockets of interest or merely on surface sites: this is the key information for structure-based design and for enabling synthesis of follow-up molecules. Extensive streamlining of the screening experiment at XChem has engendered a very active user program that is generating large amounts of data: in 2017, 36 academic and industry groups generated 35,000 datasets of uniquely soaked crystals. It has also generated a large number of learnings concerning the main remaining bottleneck, namely, obtaining a suitable crystal system that will support a successful fragment screen. Here we discuss the practicalities of generating screen-ready crystals that have useful electron density maps, and how to ensure they will be successfully reproduced and usable at a facility outside the home lab.	Oct 2018
I04-1-Macromolecular Crystallography (fixed wavelength)	An expanded allosteric network in PTP1B by multitemperature crystallography, fragment screening, and covalent tethering Daniel A. Keedy , Zachary B. Hill , Justin T. Biel , Emily Kang , T. Justin Rettenmaier , Jose Brandao-neto , Nicholas M. Pearce , Frank Von Delft , James A Wells , James Fraser Elife 7 DOI: 10.7554/eLife.36307 Diamond Proposal Number(s): [15751] Open Access Show Abstract ▼ Abstract: Allostery is an inherent feature of proteins, but it remains challenging to reveal the mechanisms by which allosteric signals propagate. A clearer understanding of this intrinsic circuitry would afford new opportunities to modulate protein function. Here we have identified allosteric sites in protein tyrosine phosphatase 1B (PTP1B) by combining multiple-temperature X-ray crystallography experiments and structure determination from hundreds of individual small-molecule fragment soaks. New modeling approaches reveal 'hidden' low-occupancy conformational states for protein and ligands. Our results converge on allosteric sites that are conformationally coupled to the active-site WPD loop and are hotspots for fragment binding. Targeting one of these sites with covalently tethered molecules or mutations allosterically inhibits enzyme activity. Overall, this work demonstrates how the ensemble nature of macromolecular structure, revealed here by multitemperature crystallography, can elucidate allosteric mechanisms and open new doors for long-range control of protein function.	Jun 2018
I04-1-Macromolecular Crystallography (fixed wavelength)	Identifying small molecule binding sites for epigenetic proteins at domain-domain interfaces Paul Brennan , David Bowkett , Romain Talon , Cynthia Tallant , Chris Schofield , Gordon Bruton , Frank Von Delft , Stefan Knapp Chemmedchem DOI: 10.1002/cmdc.201800030 Show Abstract ▼ Abstract: Epigenetics is of rapidly growing field in drug discovery. Of particular interest is the role of post‐translational modifications to histone and the proteins that read, write, and erase such modifications. The development of inhibitors for reader domains has focused on single domains. One of the major difficulties of designing inhibitors for reader domains, is that with the notable exception of bromodomains, they tend not to possess a well enclosed binding site amenable to small molecule inhibition. As many of the proteins in epigenetic regulation have multiple domains there are opportunities for designing inhibitors that bind at a domain‐domain interface which provide a more suitable interaction pocket. Examination of X‐ray structures of multiple domains involved in recognizing and modifying post‐translational histone marks using the SiteMap algorithm identified potential binding sites at domain‐domain interfaces. For the tandem plant homeodomain‐bromodomain of SP100C, a potential inter‐domain site identified computationally was validated experimentally by the discovery of ligands by X‐ray crystallographic fragment screening.	Mar 2018
	CCP 4 i 2: the new graphical user interface to the CCP 4 program suite Liz Potterton , Jon Agirre , Charles Ballard , Kevin Cowtan , Eleanor Dodson , Phil R. Evans , Huw T. Jenkins , Ronan Keegan , Eugene Krissinel , Kyle Stevenson , Andrey Lebedev , Stuart J. Mcnicholas , Robert A. Nicholls , Martin Noble , Navraj S. Pannu , Christian Roth , George Sheldrick , Pavol Skubak , Johan Turkenburg , Ville Uski , Frank Von Delft , David Waterman , Keith Wilson , Martyn Winn , Marcin Wojdyr Acta Crystallographica Section D Structural Biology 74, 68 - 84 DOI: 10.1107/S2059798317016035 Open Access Show Abstract ▼ Abstract: The CCP4 (Collaborative Computational Project, Number 4) software suite for macromolecular structure determination by X-ray crystallography groups brings together many programs and libraries that, by means of well established conventions, interoperate effectively without adhering to strict design guidelines. Because of this inherent flexibility, users are often presented with diverse, even divergent, choices for solving every type of problem. Recently, CCP4 introduced CCP4i2, a modern graphical interface designed to help structural biologists to navigate the process of structure determination, with an emphasis on pipelining and the streamlined presentation of results. In addition, CCP4i2 provides a framework for writing structure-solution scripts that can be built up incrementally to create increasingly automatic procedures.	Feb 2018
I03-Macromolecular Crystallography I04-1-Macromolecular Crystallography (fixed wavelength)	Tuning microtubule dynamics to enhance cancer therapy by modulating FER-mediated CRMP2 phosphorylation Yiyan Zheng , Ritika Sethi , Lingegowda S. Mangala , Charlotte Taylor , Juliet Goldsmith , Ming Wang , Kenta Masuda , Mohammad Karaminejadranjbar , David Mannion , Fabrizio Miranda , Sandra Herrero-gonzalez , Karin Hellner , Fiona Chen , Abdulkhaliq Alsaadi , Ashwag Albukhari , Donatien Chedom Fotso , Christopher Yau , Dahai Jiang , Sunila Pradeep , Cristian Rodriguez-aguayo , Gabriel Lopez-berestein , Stefan Knapp , Nathanael S. Gray , Leticia Campo , Kevin A. Myers , Sunanda Dhar , David Ferguson , Robert C. Bast , Anil K. Sood , Frank Von Delft , Ahmed Ashour Ahmed Nature Communications 9 DOI: 10.1038/s41467-017-02811-7 Open Access Show Abstract ▼ Abstract: Though used widely in cancer therapy, paclitaxel only elicits a response in a fraction of patients. A strong determinant of paclitaxel tumor response is the state of microtubule dynamic instability. However, whether the manipulation of this physiological process can be controlled to enhance paclitaxel response has not been tested. Here, we show a previously unrecognized role of the microtubule-associated protein CRMP2 in inducing microtubule bundling through its carboxy terminus. This activity is significantly decreased when the FER tyrosine kinase phosphorylates CRMP2 at Y479 and Y499. The crystal structures of wild-type CRMP2 and CRMP2-Y479E reveal how mimicking phosphorylation prevents tetramerization of CRMP2. Depletion of FER or reducing its catalytic activity using sub-therapeutic doses of inhibitors increases paclitaxel-induced microtubule stability and cytotoxicity in ovarian cancer cells and in vivo. This work provides a rationale for inhibiting FER-mediated CRMP2 phosphorylation to enhance paclitaxel on-target activity for cancer therapy.	Feb 2018

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