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Journal articleThinon E, Mann D, Tate EW, 2012,
Targeting N-myristoyl transferase-1 in cancer using peptide microarrays, Journal of Peptide Science, Vol: 18, Pages: S75-S75, ISSN: 1099-1387
Conference paperKonitsiotis A, Chang S, Masumoto N, et al., 2012,
Journal articlePrice HP, Hodgkinson MR, Wright MH, et al., 2012,
A role for the vesicle-associated tubulin binding protein ARL6 (BBS3) in flagellum extension in Trypanosoma brucei, Biochimica et Biophysica Acta-Molecular Cell Research, Vol: 1823, Pages: 1178-1191, ISSN: 0167-4889
The small GTPase Arl6 is implicated in the ciliopathic human genetic disorder Bardet–Biedl syndrome, actingat primary cilia in recruitment of the octomeric BBSome complex, which is required for specific traffickingevents to and from the cilium in eukaryotes. Here we describe functional characterisation of Arl6 in the flagellatedmodel eukaryote Trypanosoma brucei, which requires motility for viability. Unlike human Arl6 whichhas a ciliary localisation, TbARL6 is associated with electron-dense vesicles throughout the cell body followingco-translational modification by N-myristoylation. Similar to the related protein ARL-3A in T. brucei, modulationof expression of ARL6 by RNA interference does not prevent motility but causes a significant reductionin flagellum length. Tubulin is identified as an ARL6 interacting partner, suggesting that ARL6 may act as ananchor between vesicles and cytoplasmic microtubules. We provide evidence that the interaction betweenARL6 and the BBSome is conserved in unicellular eukaryotes. Overexpression of BBS1 leads to translocationof endogenous ARL6 to the site of exogenous BBS1 at the flagellar pocket. Furthermore, a combination ofBBS1 overexpression and ARL6 RNAi has a synergistic inhibitory effect on cell growth. Our findings indicatethat ARL6 in trypanosomes contributes to flagellum biogenesis, most likely through an interaction with theBBSome
Journal articleBell AS, Mills JE, Williams GP, et al., 2012,
Selective Inhibitors of Protozoan Protein N-myristoyltransferases as Starting Points for Tropical Disease Medicinal Chemistry Programs, PLOS Neglected Tropical Diseases, Vol: 6, ISSN: 1935-2735
Inhibition of N-myristoyltransferase has been validated pre-clinically as a target for the treatment of fungal andtrypanosome infections, using species-specific inhibitors. In order to identify inhibitors of protozoan NMTs, we chose toscreen a diverse subset of the Pfizer corporate collection against Plasmodium falciparum and Leishmania donovani NMTs.Primary screening hits against either enzyme were tested for selectivity over both human NMT isoforms (Hs1 and Hs2) andfor broad-spectrum anti-protozoan activity against the NMT from Trypanosoma brucei. Analysis of the screening results hasshown that structure-activity relationships (SAR) for Leishmania NMT are divergent from all other NMTs tested, a finding notpredicted by sequence similarity calculations, resulting in the identification of four novel series of Leishmania-selective NMTinhibitors. We found a strong overlap between the SARs for Plasmodium NMT and both human NMTs, suggesting thatachieving an appropriate selectivity profile will be more challenging. However, we did discover two novel series withselectivity for Plasmodium NMT over the other NMT orthologues in this study, and an additional two structurally distinctseries with selectivity over Leishmania NMT. We believe that release of results from this study into the public domain willaccelerate the discovery of NMT inhibitors to treat malaria and leishmaniasis. Our screening initiative is another example ofhow a tripartite partnership involving pharmaceutical industries, academic institutions and governmental/nongovernmentalorganisations such as Medical Research Council and Wellcome Trust can stimulate research for neglecteddiseases
Journal articleGoncalves V, Brannigan JA, Whalley D, et al., 2012,
Discovery of Plasmodium vivax N-Myristoyltransferase Inhibitors: Screening, Synthesis, and Structural Characterization of their Binding Mode, Journal of Medicinal Chemistry, Vol: 55, Pages: 3578-3582, ISSN: 0022-2623
N-Myristoyltransferase (NMT) is a prospective drug target against parasitic protozoa. Herein we report the successful discovery of a series of Plasmodium vivax NMT inhibitors by high-throughput screening. A high-resolution crystal structure of the hit compound in complex with NMT was obtained, allowing understanding of its novel binding mode. A set of analogues was designed and tested to define the chemical groups relevant for activity and selectivity.
Conference paperBradshaw RT, Aronica PGA, Tate EW, et al., 2012,
Developing mutational locally enhanced sampling (MULES) for predicting relative binding free energies at protein-protein interfaces, 11th International Biorelated Polymer Symposium / 243rd National Spring Meeting of the American-Chemical-Society (ACS), Publisher: AMER CHEMICAL SOC, ISSN: 0065-7727
Journal articleGoncalves V, Brannigan JA, Thinon E, et al., 2012,
N-myristoylation is the irreversible attachment of a C14 fatty acid, myristic acid, to the N-terminal glycine of a protein via formation of an amide bond. This modification is catalyzed by myristoyl–coenzyme A (CoA):protein N-myristoyltransferase (NMT), an enzyme ubiquitous in eukaryotes that is up-regulated in several cancers. Here we report a sensitive fluorescence-based assay to study the enzymatic activity of human NMT1 and NMT2 based on detection of CoA by 7-diethylamino-3-(4-maleimido-phenyl)-4-methylcoumarin. We also describe expression and characterization of NMT1 and NMT2 and assay validation with small molecule inhibitors. This assay should be broadly applicable to NMTs from a range of organisms.
Journal articleBroncel M, Serwa RA, Tate EW, 2012,
agging protein AMPylation: A new chemical reporter for AMPylation, recently identified as a key post-translational modification during bacterial infection, is a robust tool for detecting and identifying AMPylated proteins in vitro.
Journal articleDang THT, Fagan RP, Fairweather NF, et al., 2012,
Clostridium difficile, a leading cause of hospital-acquired bacterial infection, is coated in a dense surface layer (S-layer) that is thought to provide both physicochemical protection and a scaffold for host-pathogen interactions. The key structural components of the S-layer are two proteins derived from a polypeptide precursor, SlpA, via proteolytic cleavage by the protease Cwp84. Here, we report the design, synthesis and in vivo characterization of a panel of protease inhibitors and activity-based probes (ABPs) designed to target S-layer processing in live C. difficile cells. Inhibitors based on substrate-mimetic peptides bearing a C-terminal Michael acceptor warhead were found to be promising candidates for further development.
Journal articleHeal WP, Tate EW, 2012,
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