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18 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 I24-Microfocus Macromolecular Crystallography	Scaffold-hopping identifies furano[2,3-d]pyrimidine amides as potent Notum inhibitors Benjamin N. Atkinson , David Steadman , William Mahy , Yuguang Zhao , James Sipthorp , Elliott D. Bayle , Fredrik Svensson , George Papageorgiou , Fiona Jeganathan , Sarah Frew , Amy Monaghan , Magda Bictash , E. Yvonne Jones , Paul V. Fish Bioorganic & Medicinal Chemistry Letters DOI: 10.1016/j.bmcl.2019.126751 Diamond Proposal Number(s): [14744] Show Abstract ▼ Abstract: The carboxylesterase Notum is a key negative regulator of the Wnt signaling pathway by mediating the depalmitoleoylation of Wnt proteins. Our objective was to discover potent small molecule inhibitors of Notum suitable for exploring the regulation of Wnt signaling in the central nervous system. Scaffold-hopping from thienopyrimidine acids 1 and 2, supported by X-ray structure determination, identified 3-methylimidazolin-4-one amides 20-24 as potent inhibitors of Notum with activity across three orthogonal assay formats (biochemical, extra-cellular, occupancy). A preferred example 24 demonstrated good stability in mouse microsomes and plasma, and cell permeability in the MDCK-MDR1 assay albeit with modest P-gp mediated efflux. Pharmacokinetic studies with 24 were performed in vivo in mouse with single oral administration of 24 showing good plasma exposure and reasonable CNS penetration. We propose that 24 is a new chemical tool suitable for cellular studies to explore the fundamental biology of Notum.	Oct 2019
I04-1-Macromolecular Crystallography (fixed wavelength) I24-Microfocus Macromolecular Crystallography	Discovery of 2-phenoxyacetamides as inhibitors of the Wnt-depalmitoleating enzyme NOTUM from an X-ray fragment screen Benjamin Nicholas Atkinson , David Steadman , Yuguang Zhao , James Sipthorp , Luca Vecchia , Reinis Reinholds Ruza , Fiona Jeganathan , Georgie Lines , Sarah Frew , Amy Monaghan , Svend Kjaer , Magda Bictash , Yvonne Jones , Paul V. Fish Medchemcomm DOI: 10.1039/C9MD00096H Diamond Proposal Number(s): [16814, 14744] Open Access Show Abstract ▼ Abstract: NOTUM is a carboxylesterase that has been shown to act by mediating the O-depalmitoleoylation of Wnt proteins resulting in suppression of Wnt signaling. Here, we describe the development of NOTUM inhibitors that restore Wnt signaling for use in in vitro disease models where NOTUM over activity is an underlying cause. A crystallographic fragment screen with NOTUM identified 2-phenoxyacetamide 3 as binding in the palmitoleate pocket with modest inhibition activity (IC50 33 μM). Optimization of hit 3 by SAR studies guided by SBDD identified indazole 38 (IC50 0.032 μM) and isoquinoline 45 (IC50 0.085 μM) as potent inhibitors of NOTUM. The binding of 45 to NOTUM was rationalized through an X-ray cocrystal structure determination which showed a flipped binding orientation compared to 3. However, it was not possible to combine NOTUM inhibition activity with metabolic stability as the majority of the compounds tested were rapidly metabolized in an NADPH-independent manner.	Apr 2019
I04-1-Macromolecular Crystallography (fixed wavelength) I24-Microfocus Macromolecular Crystallography	Structure-based in silico screening identifies a potent ebolavirus inhibitor from a traditional Chinese medicine library Faraz Shaikh , Yuguang Zhao , Luis Alvarez , Maria Iliopoulou , Christopher Thomas Lohans , Christopher J. Schofield , Sergi Padilla-parra , Shirley W. I. Siu , Elizabeth Fry , Jingshan Ren , David I. Stuart Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.8b01328 Diamond Proposal Number(s): [10627] Show Abstract ▼ Abstract: Potent Ebolavirus (EBOV) inhibitors will help to curtail outbreaks such as that which occurred in 2014-16 in West Africa. EBOV has on its surface a single glycoprotein (GP) critical for viral entry and membrane fusion. Recent high resolution complexes of EBOV GP with a variety of approved drugs revealed that binding to a common cavity prevented fusion of the virus and endosomal membranes, inhibiting virus infection. We performed docking experiments, screening a database of natural compounds to identify those likely to bind at this site. Using both inhibition assays of HIV-1-derived pseudovirus cell entry and structural analyses of the complexes of the compounds with GP we show here that two of these compounds attach in the common binding cavity, out of eight tested. In both cases two molecules bind in the cavity. The two compounds are chemically similar but the tighter binder has an additional chlorine atom that forms good halogen bonds to the protein and achieves an IC50 of 50 nM, making it the most potent GP-binding EBOV inhibitor yet identified, validating our screening approach for the discovery of novel anti-viral compounds.	Feb 2019
	Unexpected mode of engagement between enterovirus 71 and its receptor SCARB2 Daming Zhou , Yuguang Zhao , Abhay Kotecha , Elizabeth E. Fry , James T. Kelly , Xiangxi Wang , Zihe Rao , David J. Rowlands , Jingshan Ren , David I. Stuart Nature Microbiology 25 DOI: 10.1038/s41564-018-0319-z Show Abstract ▼ Abstract: Enterovirus 71 (EV71) is a common cause of hand, foot and mouth disease—a disease endemic especially in the Asia-Pacific region1. Scavenger receptor class B member 2 (SCARB2) is the major receptor of EV71, as well as several other enteroviruses responsible for hand, foot and mouth disease, and plays a key role in cell entry2. The isolated structures of EV71 and SCARB2 are known3,4,5,6, but how they interact to initiate infection is not. Here, we report the EV71–SCARB2 complex structure determined at 3.4 Å resolution using cryo-electron microscopy. This reveals that SCARB2 binds EV71 on the southern rim of the canyon, rather than across the canyon, as predicted3,7,8. Helices 152–163 (α5) and 183–193 (α7) of SCARB2 and the viral protein 1 (VP1) GH and VP2 EF loops of EV71 dominate the interaction, suggesting an allosteric mechanism by which receptor binding might facilitate the low-pH uncoating of the virus in the endosome/lysosome. Remarkably, many residues within the binding footprint are not conserved across SCARB2-dependent enteroviruses; however, a conserved proline and glycine seem to be key residues. Thus, although the virus maintains antigenic variability even within the receptor-binding footprint, the identification of binding ‘hot spots’ may facilitate the design of receptor mimic therapeutics less likely to quickly generate resistance.	Dec 2018
I03-Macromolecular Crystallography I04-Macromolecular Crystallography	Structure of the Wnt signaling enhancer LYPD6 and its interactions with the Wnt co-receptor LRP6 Yuguang Zhao , Jingshan Ren , Weixian Lu , Karl Harlos , E. Yvonne Jones Febs Letters DOI: 10.1002/1873-3468.13212 Diamond Proposal Number(s): [8423] Show Abstract ▼ Abstract: LYPD6 is a Wnt signaling enhancer that promotes phosphorylation of the Wnt co‐receptor LRP6 (low‐density lipoprotein receptor‐related protein 6). It also binds the nicotinic acetylcholine receptor (nAChR). We report here the 1.25 Å resolution structure of the LYPD6 extracellular Ly6/uPAR (LU) domain and map its interaction with LRP6 by mutagenesis and surface plasmon resonance (SPR). The LYPD6LU structure reveals a “tri‐fingered protein domain” fold with the middle fingertip of the bearing a “NxI” motif, a tripeptide motif associated with LRP5/6 binding by Wnt inhibitors. Of the Ly6 protein family members, only LYPD6 has an NxI motif. Since mutations in the LYPD6 NxI motif abolish or severely reduce interaction with LRP6, our results indicate its key role in the interaction of LYPD6 with LRP6.	Aug 2018
I04-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Structures of ebolavirus glycoprotein complexes with tricyclic antidepressant and antipsychotic drugs Yuguang Zhao , Jingshan Ren , Elizabeth E. Fry , Julia Xiao , Alain R. Townsend , David I. Stuart Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.8b00350 Diamond Proposal Number(s): [10627] Show Abstract ▼ Abstract: A large number of Food and Drug Administration (FDA) approved drugs have been found to inhibit cell entry of Ebola virus (EBOV). However, since these drugs have various primary pharmacological targets their mechanisms of action against EBOV remain largely unknown. We have previously shown that six FDA approved drugs inhibit EBOV infection by interacting with and destabilizing the viral glycoprotein (GP). Here we show that the antidepressants imipramine and clomipramine, and antipsychotic drug thioridazine also directly interact with EBOV GP, and determine the mode of interaction by crystallographic analysis of the complexes. The compounds bind within the same pocket as observed for other, chemically divergent, complexes but with different binding modes. These details should be of value for the development of potent EBOV inhibitors.	May 2018
I02-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Target identification and mode of action of four chemically divergent drugs against Ebola virus infection Jingshan Ren , Yuguang Zhao , Elizabeth E. Fry , David I. Stuart Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.7b01249 Diamond Proposal Number(s): [10627] Show Abstract ▼ Abstract: Here we show that four chemically divergent approved drugs reported to inhibit Ebolavirus infection, benztropine, bepridil, paroxetine and sertraline, directly interact with the Ebolavirus glycoprotein. Binding of these drugs destabilise the protein, suggesting that this may be the mechanism of inhibition, as reported for the anticancer drug toremifene and the painkiller ibuprofen, which bind in the same large cavity on the glycoprotein. Crystal structures show that the position of binding and the mode of interaction within the pocket vary significantly between these compounds. The binding constants (Kd) determined by thermal shift assay correlate with the protein-inhibitor interactions as well as with the antiviral activities determined by virus cell entry assays, supporting the hypothesis that these drugs inhibit viral entry by binding the glycoprotein and destabilising the pre-fusion conformation. Details of the protein–inhibitor interactions of these complexes and their relation with binding affinity may facilitate the design of more potent inhibitors.	Dec 2017
I03-Macromolecular Crystallography I24-Microfocus Macromolecular Crystallography	Stereotyped antibody responses target posttranslationally modified gluten in celiac disease Omri Snir , Xi Chen , Moriah Gidoni , M. Fleur Du Pré , Yuguang Zhao , Øyvind Steinsbø , Knut E. A. Lundin , Gur Yaari , Ludvig M. Sollid Jci Insight 2 DOI: 10.1172/jci.insight.93961 Diamond Proposal Number(s): [10627] Show Abstract ▼ Abstract: The role of B cells and posttranslational modifications in pathogenesis of organ-specific immune diseases is increasingly envisioned but remains poorly understood, particularly in human disorders. In celiac disease, transglutaminase 2–modified (TG2-modified; deamidated) gluten peptides drive disease-specific T cell and B cell responses, and antibodies to deamidated gluten peptides are excellent diagnostic markers. Here, we substantiate by high-throughput sequencing of IGHV genes that antibodies to a disease-specific, deamidated, and immunodominant B cell epitope of gluten (PLQPEQPFP) have biased and stereotyped usage of IGHV3-23 and IGHV3-15 gene segments with modest somatic mutations. X-ray crystal structures of 2 prototype IGHV3-15/IGKV4-1 and IGHV3-23/IGLV4-69 antibodies reveal peptide interaction mainly via germline-encoded residues. In-depth mutational analysis showed restricted selection and substitution patterns at positions involved in antigen binding. While the IGHV3-15/IGKV4-1 antibody interacts with Glu5 and Gln6, the IGHV3-23/IGLV4-69 antibody interacts with Gln3, Pro4, Pro7, and Phe8 — residues involved in substrate recognition by TG2. Hence, both antibodies, despite different interaction with the epitope, recognize signatures of TG2 processing that facilitates B cell presentation of deamidated gluten peptides to T cells, thereby providing a molecular framework for the generation of these clinically important antibodies. The study provides essential insight into the pathogenic mechanism of celiac disease.	Sep 2017
I03-Macromolecular Crystallography	Ligand-induced conformational change of Insulin-regulated aminopeptidase: insights on catalytic mechanism and active site plasticity Anastasia Mpakali , Emmanuel Saridakis , Karl Harlos , Yuguang Zhao , Paraskevi Kokkala , Dimitris Georgiadis , Petros Giastas , Athanasios Papakyriakou , Efstratios Stratikos Journal Of Medicinal Chemistry DOI: 10.1021/acs.jmedchem.6b01890 Diamond Proposal Number(s): [10627] Show Abstract ▼ Abstract: Insulin-regulated aminopeptidase (IRAP) is an enzyme with several important biological functions that is known to process a large variety of different peptidic substrates although the mechanism behind this wide specificity is not clearly understood. We describe a crystal structure of IRAP in complex with a recently developed bioactive and selective inhibitor at 2.53 Å resolution. In the presence of this inhibitor the enzyme adopts a novel conformation in which domains II and IV are juxtaposed, forming a hollow structure that excludes external solvent access to the catalytic center. A loop adjacent to the enzyme’s GAMEN motif undergoes structural reconfiguration, allowing the accommodation of bulky inhibitor side-chains. Atomic interactions between the inhibitor and IRAP that are unique to this conformation can explain the strong selectivity compared to homologous aminopeptidases ERAP1 and ERAP2. This conformation provides insight on IRAP’s catalytic cycle and reveals significant active site plasticity that may underlie its substrate permissiveness.	Mar 2017
I03-Macromolecular Crystallography I04-Macromolecular Crystallography	Structure of the Dual-Mode Wnt Regulator Kremen1 and Insight into Ternary Complex Formation with LRP6 and Dickkopf Matthias Zebisch , Verity a. Jackson , Yuguang Zhao , E. yvonne Jones Structure 24, 1599 - 1605 DOI: 10.1016/j.str.2016.06.020 Diamond Proposal Number(s): [8423, 10627] Open Access Show Abstract ▼ Abstract: Kremen 1 and 2 have been identified as co-receptors for Dickkopf (Dkk) proteins, hallmark secreted antagonists of canonical Wnt signaling. We present here three crystal structures of the ectodomain of human Kremen1 (KRM1ECD) at resolutions between 1.9 and 3.2 Å. KRM1ECD emerges as a rigid molecule with tight interactions stabilizing a triangular arrangement of its Kringle, WSC, and CUB structural domains. The structures reveal an unpredicted homology of the WSC domain to hepatocyte growth factor. We further report the general architecture of the ternary complex formed by the Wnt co-receptor Lrp5/6, Dkk, and Krm, determined from a low-resolution complex crystal structure between β-propeller/EGF repeats (PE) 3 and 4 of the Wnt co-receptor LRP6 (LRP6PE3PE4), the cysteine-rich domain 2 (CRD2) of DKK1, and KRM1ECD. DKK1CRD2 is sandwiched between LRP6PE3 and KRM1Kringle-WSC. Modeling studies supported by surface plasmon resonance suggest a direct interaction site between Krm1CUB and Lrp6PE2.	Sep 2016

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