25 results on '"Sim, Edith"'
Search Results
2. Arylamine N-Acetyltransferases in Normal and Abnormal Embryonic Development
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Stanley, Lesley A., primary and Sim, Edith, additional
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- 2018
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3. Fusarium verticillioides NAT1 ( FDB2 ) N ‐malonyltransferase is structurally, functionally and phylogenetically distinct from its N ‐acetyltransferase ( NAT ) homologues
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Karagianni, Eleni‐Pavlina, primary, Kontomina, Evanthia, additional, Lowe, Edward D., additional, Athanasopoulos, Konstantinos, additional, Papanikolaou, Georgia, additional, Garefalaki, Vasiliki, additional, Kotseli, Varvara, additional, Zaliou, Sofia, additional, Grimaud, Tom, additional, Arvaniti, Konstantina, additional, Tsatiri, Maria‐Aggeliki, additional, Fakis, Giannoulis, additional, Glenn, Anthony E., additional, Roversi, Pietro, additional, Abuhammad, Areej, additional, Ryan, Ali, additional, Sim, Robert B., additional, Sim, Edith, additional, and Boukouvala, Sotiria, additional
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- 2022
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4. Recent progress in N-acetyltransferase research: 7th international workshop on N-acetyltransferases (NAT): workshop report
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Lichter, Jutta, Golka, Klaus, Sim, Edith, and Blömeke, Brunhilde
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- 2017
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5. Arylamine N-Acetyltransferases in Health and Disease
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Laurieri, Nicola, primary and Sim, Edith, additional
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- 2017
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6. Fusarium verticillioidesNAT1 (FDB2) N‐malonyltransferase is structurally, functionally and phylogenetically distinct from its N‐acetyltransferase (NAT) homologues.
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Karagianni, Eleni‐Pavlina, Kontomina, Evanthia, Lowe, Edward D., Athanasopoulos, Konstantinos, Papanikolaou, Georgia, Garefalaki, Vasiliki, Kotseli, Varvara, Zaliou, Sofia, Grimaud, Tom, Arvaniti, Konstantina, Tsatiri, Maria‐Aggeliki, Fakis, Giannoulis, Glenn, Anthony E., Roversi, Pietro, Abuhammad, Areej, Ryan, Ali, Sim, Robert B., Sim, Edith, and Boukouvala, Sotiria
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GIBBERELLA fujikuroi ,FUSARIUM ,ISOENZYMES ,MONOMERS ,HYDROGEN bonding ,ACETYLCOENZYME A ,METABOLIC detoxification ,MYCOTOXINS - Abstract
Fusarium endophytes damage cereal crops and contaminate produce with mycotoxins. Those fungi overcome the main chemical defence of host via detoxification by a malonyl‐CoA‐dependent enzyme homologous to xenobiotic metabolizing arylamine N‐acetyltransferase (NAT). In Fusarium verticillioides (teleomorph Gibberella moniliformis, GIBMO), this N‐malonyltransferase activity is attributed to (GIBMO)NAT1, and the fungus has two additional isoenzymes, (GIBMO)NAT3 (N‐acetyltransferase) and (GIBMO)NAT2 (unknown function). We present the crystallographic structure of (GIBMO)NAT1, also modelling other fungal NAT homologues. Monomeric (GIBMO)NAT1 is distinctive, with access to the catalytic core through two "tunnel‐like" entries separated by a "bridge‐like" helix. In the quaternary arrangement, (GIBMO)NAT1 monomers interact in pairs along an extensive interface whereby one entry of each monomer is covered by the N‐terminus of the other monomer. Although monomeric (GIBMO)NAT1 apparently accommodates acetyl‐CoA better than malonyl‐CoA, dimerization changes the active site to allow malonyl‐CoA to reach the catalytic triad (Cys110, His158 and Asp173) via the single uncovered entry, and anchor its terminal carboxyl‐group via hydrogen bonds to Arg109, Asn157 and Thr261. Lacking a terminal carboxyl‐group, acetyl‐CoA cannot form such stabilizing interactions, while longer acyl‐CoAs enter the active site but cannot reach catalytic Cys. Other NAT isoenzymes lack such structural features, with (GIBMO)NAT3 resembling bacterial NATs and (GIBMO)NAT2 adopting a structure intermediate between (GIBMO)NAT1 and (GIBMO)NAT3. Biochemical assays confirmed differential donor substrate preference of (GIBMO)NAT isoenzymes, with phylogenetic analysis demonstrating evolutionary separation. Given the role of (GIBMO)NAT1 in enhancing Fusarium pathogenicity, unravelling the structure and function of this enzyme may benefit research into more targeted strategies for pathogen control. [ABSTRACT FROM AUTHOR]
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- 2023
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7. Comparative analysis of xenobiotic metabolising N-acetyltransferases from ten non-human primates as in vitro models of human homologues
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Tsirka, Theodora, Konstantopoulou, Maria, Sabbagh, Audrey, Crouau-Roy, Brigitte, Ryan, Ali, Sim, Edith, Boukouvala, Sotiria, and Fakis, Giannoulis
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- 2018
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8. Investigation of the mycobacterial enzyme HsaD as a potential novel target for anti-tubercular agents using a fragment-based drug design approach
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Ryan, Ali, Polycarpou, Elena, Lack, Nathan A., Evangelopoulos, Dimitrios, Sieg, Christian, Halman, Alice, Bhakta, Sanjib, Eleftheriadou, Olga, McHugh, Timothy D., Keany, Sebastian, Lowe, Edward D., Ballet, Romain, Abuhammad, Areej, Jacobs Jr., William R., Ciulli, Alessio, and Sim, Edith
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pharmacy ,chemistry ,biological - Published
- 2017
9. FRONT MATTER
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Laurieri, Nicola, primary and Sim, Edith, additional
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- 2018
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10. BACK MATTER
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Laurieri, Nicola, primary and Sim, Edith, additional
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- 2018
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11. Identification of novel members of the bacterial azoreductase family in 'Pseudomonas aeruginosa'
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Crescente, Vincenzo, Holland, Sinead, Kashyap, Sapna, Polycarpou, Elena, Sim, Edith, and Ryan, Ali
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biological - Abstract
Azoreductases are a family of diverse enzymes found in many pathogenic bacteria as well as distant homologues being present in eukarya. In addition to having azoreductase activity these enzymes are also suggested to have NAD(P)H quinone oxidoreductase activity which leads to a proposed role in plant pathogenesis. Azoreductases have also been suggested to play role in the mammalian pathogenesis of Pseudomonas aeruginosa. In view of the importance of P. aeruginosa as a pathogen, we therefore characterised recombinant enzymes following expression of a group of putative azoreductase genes from P. aeruginosa expressed in Escherichia coli . The enzymes include members of the "Arsenic resistance protein H" (ArsH), "tryptophan repressor binding protein A" (WrbA), "modulator of drug activity B" (MdaB) and YieF families. The ArsH, MdaB and YieF family members all show azoreductase and NAD(P)H quinone oxidoreductase activities. In contrast, WrbA is the first enzyme to show NAD(P)H quinone oxidoreductase activity but does not reduce any of the 11 azo compounds tested under a wide range of conditions. These studies will allow further investigation of the possible role of these enzymes in the pathogenesis of P. aeruginosa .
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- 2016
12. Arylamine N-Acetyltransferases
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Boukouvala, Sotiria, Fakis, Giannoulis, Stavrakaki, Ioanna, Polycarpou, Elena, Laurieri, Nicola, and Sim, Edith
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- 2015
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13. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway
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Fineran, Paul, primary, Lloyd-Evans, Emyr, additional, Lack, Nathan A., additional, Platt, Nick, additional, Davis, Lianne C., additional, Morgan, Anthony J., additional, Höglinger, Doris, additional, Tatituri, Raju Venkata V., additional, Clark, Simon, additional, Williams, Ian M., additional, Tynan, Patricia, additional, Al Eisa, Nada, additional, Nazarova, Evgeniya, additional, Williams, Ann, additional, Galione, Antony, additional, Ory, Daniel S., additional, Besra, Gurdyal S., additional, Russell, David G., additional, Brenner, Michael B., additional, Sim, Edith, additional, and Platt, Frances M., additional
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- 2017
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14. Drug metabolism and antibiotic resistance in micro-organisms
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Sim, Edith, primary and Ryan, Ali, additional
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- 2017
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15. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway
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Fineran, Paul, primary, Lloyd-Evans, Emyr, additional, Lack, Nathan A., additional, Platt, Nick, additional, Davis, Lianne C., additional, Morgan, Anthony J., additional, Höglinger, Doris, additional, Tatituri, Raju Venkata V., additional, Clark, Simon, additional, Williams, Ian M., additional, Tynan, Patricia, additional, Al Eisa, Nada, additional, Nazarova, Evgeniya, additional, Williams, Ann, additional, Galione, Antony, additional, Ory, Daniel S., additional, Besra, Gurdyal S., additional, Russell, David G., additional, Brenner, Michael B., additional, Sim, Edith, additional, and Platt, Frances M., additional
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- 2016
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16. Homologues of xenobiotic metabolizing N-acetyltransferases in plant-associated fungi: Novel functions for an old enzyme family
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Karagianni, Eleni P., primary, Kontomina, Evanthia, additional, Davis, Britton, additional, Kotseli, Barbara, additional, Tsirka, Theodora, additional, Garefalaki, Vasiliki, additional, Sim, Edith, additional, Glenn, Anthony E., additional, and Boukouvala, Sotiria, additional
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- 2015
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17. Characterisation of a putative AraC transcriptional regulator from Mycobacterium smegmatis
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Evangelopoulos, Dimitrios, primary, Gupta, Antima, additional, Lack, Nathan A., additional, Maitra, Arundhati, additional, ten Bokum, Annemieke M.C., additional, Kendall, Sharon, additional, Sim, Edith, additional, and Bhakta, Sanjib, additional
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- 2014
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18. Differences between murine arylamine N-acetyltransferase type 1 and human arylamine N-acetyltransferase type 2 defined by substrate specificity and inhibitor binding
- Author
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Laurieri, Nicola, primary, Kawamura, Akane, additional, Westwood, Isaac M, additional, Varney, Amy, additional, Morris, Elizabeth, additional, Russell, Angela J, additional, Stanley, Lesley A, additional, and Sim, Edith, additional
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- 2014
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19. Exploration of Piperidinols as Potential Antitubercular Agents
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Abuhammad, Areej, primary, Fullam, Elizabeth, additional, Bhakta, Sanjib, additional, Russell, Angela, additional, Morris, Garrett, additional, Finn, Paul, additional, and Sim, Edith, additional
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- 2014
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20. Azoreductases : genes and proteins in Pseudomonas aeruginosa
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Crescente, Vincenzo and Sim, Edith
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616.9 - Abstract
Pseudomonas aeruginosa is one of the primary causes of opportunistic infections in humans and it is associated with both acute and chronic infections in immunocompromised individuals. This bacterium is extremely resistant to many antibiotics, making the treatments against this pathogen often unsuccessful. Azoreductases, a family of enzymes involved in the reduction of azo compounds and quinones, are found in many bacterial species including P. aeruginosa. Although the enzymatic activity of three of these enzymes has been extensively characterized, their physiological role remains unclear. In this study, the enzymatic activity as well as the effect on physiological processes such as swarming motility, biofilm formation and antibiotic resistance of known and putative azoreductase proteins from P. aeruginosa PAO1 have been investigated. Five putative azoreductase genes from P. aeruginosa PAO1 (pa0949, pa1204, pa2280, pa2580 and pa4975) were cloned and four of these (pa0949, pa1204, pa2280 and pa2580) were over expressed in E. coli strains. Recombinant proteins were purified and biochemically characterized showing the presence of FAD bound to PA1204 and PA2580 proteins. Enzymatic reaction conditions were established for each protein by determining the preferred cofactor and reductant (flavin and NAD(P)H) used by each protein. Higher reduction rates were obtained using FAD for PA1204 and PA2580, and FMN for PA0949 and PA2280, whereas NADPH was always the preferred reductant for all of the enzymes tested. Substrate specificity studies performed with azo compounds and quinones showed that PA1204, PA2280 and PA2580 recombinant proteins can reduce both classes of substrate, with higher reduction rates with quinones, whereas the recombinant PA0949 protein showed to reduce only quinone substrates. Investigation of the role of azoreductase genes on P. aeruginosa PAO1 motility, biofilm formation and antibiotic resistance was conducted using single gene transposon mutants for each of the genes paazor1, paazor2, paazor3, pa0949, pa1204, pa2280, pa2580 and pa4975. Motility analysis demonstrated greater swarming for all mutants tested compared with wild type, although both wild type and mutants showed the same growth rate. Similarly, all mutants showed higher biofilm production compared with the wild type in a short term period (24 hours), whereas no differences were observed after a longer biofilm production period (48 hours). The MICs and MBCs of several antibiotics were determined, showing that, in presence of fluoroquinolones, P. aeruginosa PAO1 azoreductase mutants exhibit higher growth inhibition (up to 127-fold) and reduced survival (up to 7 fold) compared with wild type. This suggests that azoreductase genes (or gene products) may be involved in the P. aeruginosa PAO1 resistance to antibiotic treatments, and in particular to fluoroquinolones. The findings prove that the proteins PA0949, PA1204, PA2280 and PA2580 have similar features and enzymatic functions to the already characterized paAzoR1-3 from P. aeruginosa PAO1. Therefore, these can be included in the family of azo- and quinone- oxidoreductase enzymes. The data presented here on the antibiotic resistance strongly suggest a role for azoreductase gene products in antimicrobial resistance in P. aeruginosa. These original findings provide a springboard for further investigation of azoreductases as novel targets for antimicrobial agents for this pathogen.
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- 2015
21. Small molecule colorimetric and fluorescent probes for specific protein detection
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Egleton, James Edward, Russell, Angela, and Sim, Edith
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547 ,Organic chemistry ,Chemical biology ,Enzymes ,Pharmacology ,colorimetric probe ,fluorescent probe ,protein detection ,arylamine N-acetyltransferase ,breast cancer - Abstract
This thesis describes the design, synthesis, analysis, mechanistic evaluation and optimisation of small molecule probes for the specific detection of proteins, focusing on the target protein human arylamine N-acetyltransferase type 1 (HUMAN(NAT1)) and its murine homologue, mouse arylamine N-acetyltransferase type 2 (MOUSE(NAT2)). The HUMAN(NAT1) gene is reported to be one of the most highly overexpressed genes in estrogen-receptor-positive (ER+) breast tumours, leading to its potential use as both a novel diagnostic biomarker and a novel therapeutic target for this disease. Chapter 1 reviews the literature on optical methods for the specific detection of a protein target, exploring strategies both based on biosensors and on chemical probes, before introducing the arylamine N-acetyltransferases as a family of enzymes. In Chapter 2, a family of naphthoquinone inhibitors of HUMAN(NAT1) are introduced, which undergo a colour change from red to blue upon binding specifically to the enzyme. The mechanism of this colour change, a proton transfer-mediated process, is discussed via the synthesis, pharmacological and colorimetric evaluation of close analogues of the hit compound lacking a key acidic sulfonamide-NH proton. During these studies, it was found that direct O-methylation of a sulfonamide is possible under certain conditions; such a reaction has not previously been reported. Furthermore, upon heating in polar solvents the O-methylated sulfonamide was observed to undergo rearrangement, and the mechanism of this process is investigated via NMR and kinetic studies. In Chapter 3, the design, synthesis and evaluation of HUMAN(NAT1) inhibitors with improved pharmacological and colorimetric profiles over the initial hit are described. From this optimisation, structure-activity relationships and an in silico model of interactions between the inhibitors and enzyme are evaluated. Testing of these compounds in cellular environments, however, exposes some limitations of this approach, notably the lack of sensitivity of the probes when dosed at low concentrations in cellular samples. In order to overcome this limitation, in Chapter 4 fluorescent analogues of the hit compound are designed and synthesised. Initial compounds developed in this series possess promising properties, but each compound generated suffers from either a low fluorescent intensity, lack of a pH-dependent switch in fluorescence or a low fluorescence excitation wavelength, which overlaps with those of tryptophan or tyrosine residues in proteins. Insights into the mechanism of molecular fluorescence and application of some simple quantum mechanical principles, however, lead to the design of a species which possesses all the required properties. The fluorescent emission intensity of this probe correlates linearly with [MOUSE(NAT2)] in E. coli cell extracts, and can quantify as little as 0.64% MOUSE(NAT2) in the samples; furthermore, the probe is capable of unambiguously detecting HUMAN(NAT1) within a cell extract from the ER+ breast cancer cell line ZR-75-1; future work on this probe may therefore enable its clinical use in improved early diagnosis of breast tumours. This study also represents, to the best of our knowledge, the first ever example of a small molecule, non-covalent probe capable of quantifying the concentration of a target protein in cellular extracts. In Chapter 5, the series of naphthoquinone probes is further optimised in order to study the roles of HUMAN(NAT1) in a cellular environment. Firstly, structure-activity relationships are utilised to design inhibitors with improved physical properties such as aqueous solubility and cell membrane permeability, in order to test the effect of HUMAN(NAT1) inhibitors in tumour cell models, which could have implications for the future use of a HUMAN(NAT1) inhibitor as a therapeutic agent in oncology. Secondly, the effect of the cofactor folic acid on the function and activity of HUMAN(NAT1) is explored. Finally, in Chapter 6, the conclusions of this study are outlined and a hypothesis as to how the concepts developed in this thesis might be applied to alternative, more ubiquitous biological targets is discussed, paving the way for future investigations.
- Published
- 2015
22. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway
- Author
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Simon Clark, Anthony J. Morgan, Raju V. V. Tatituri, Emyr Lloyd-Evans, Lianne C. Davis, Frances M. Platt, Edith Sim, Nick Platt, David G. Russell, Doris Höglinger, Nathan A. Lack, Evgeniya V. Nazarova, Ann Williams, Paul Fineran, Nada Al Eisa, Patricia W. Tynan, Gurdyal S. Besra, Daniel S. Ory, Antony Galione, Ian M. Williams, Michael B. Brenner, Lack, Nathan Alan (ORCID 0000-0001-7399-5844 & YÖK ID 120842), Fineran, Paul, Lloyd-Evans, Emyr, Platt, Nick, Davis, Lianne C., Morgan, Anthony J., Höglinger, Doris, Tatituri, Raju Venkata Veera, Clark, Simon O., Williams, Ian M., Tynan, Patricia W., Eisa, Nada Al, Nazarova, Evgeniya V., Williams, Ann I.O., Galione, Antony G., Ory, Daniel S., Besra, Gurdyal Singh, Russell, David G., Brenner, Michael B., Sim, Edith, Platt, Frances M., School of Medicine, and Department of Pharmacology
- Subjects
0301 basic medicine ,Programmed cell death ,Endocytic cycle ,Cell ,Medicine (miscellaneous) ,Tuberculosis ,Niemann-pick Disease Type C ,Lysosomal Storage Diseases ,Lysosomal calcium ,Bioinformatics ,General Biochemistry, Genetics and Molecular Biology ,Niemann-Pick Disease Type C ,03 medical and health sciences ,0302 clinical medicine ,Lysosome ,Lysosomal storage disease ,Genetics of the Immune System ,Medicine ,Pharmacology ,Niemann–Pick disease, type C ,business.industry ,Articles ,medicine.disease ,Cell biology ,3. Good health ,medicine.anatomical_structure ,030104 developmental biology ,Respiratory Infections ,NPC1 ,Lysosomal Calcium ,business ,Intracellular ,030217 neurology & neurosurgery ,Research Article - Abstract
Background. Tuberculosis remains a major global health concern. The ability to prevent phagosome-lysosome fusion is a key mechanism by which intracellular mycobacteria, including Mycobacterium tuberculosis, achieve long-term persistence within host cells. The mechanisms underpinning this key intracellular pro-survival strategy remain incompletely understood. Host macrophages infected with intracellular mycobacteria share phenotypic similarities with cells taken from patients suffering from Niemann-Pick Disease Type C (NPC), a rare lysosomal storage disease in which endocytic trafficking defects and lipid accumulation within the lysosome lead to cell dysfunction and cell death. We investigated whether these shared phenotypes reflected an underlying mechanistic connection between mycobacterial intracellular persistence and the host cell pathway dysfunctional in NPC. Methods. The induction of NPC phenotypes in macrophages from wild-type mice or obtained from healthy human donors was assessed via infection with mycobacteria and subsequent measurement of lipid levels and intracellular calcium homeostasis. The effect of NPC therapeutics on intracellular mycobacterial load was also assessed. Results. Macrophages infected with intracellular mycobacteria phenocopied NPC cells, exhibiting accumulation of multiple lipid types, reduced lysosomal Ca2+ levels, and defects in intracellular trafficking. These NPC phenotypes could also be induced using only lipids/glycomycolates from the mycobacterial cell wall. These data suggest that intracellular mycobacteria inhibit the NPC pathway, likely via inhibition of the NPC1 protein, and subsequently induce altered acidic store Ca2+ homeostasis. Reduced lysosomal calcium levels may provide a mechanistic explanation for the reduced levels of phagosome-lysosome fusion in mycobacterial infection. Treatments capable of correcting defects in NPC mutant cells via modulation of host cell calcium were of benefit in promoting clearance of mycobacteria from infected host cells. Conclusion. These findings provide a novel mechanistic explanation for mycobacterial intracellular persistence, and suggest that targeting interactions between the mycobacteria and host cell pathways may provide a novel avenue for development of anti-TB therapies., Wellcome Trust; National Institutes of Health; Natural Sciences and Engineering Research Council of Canada; Rosetrees Trust; Research Councils UK
- Published
- 2017
23. Fusarium verticillioides NAT1 (FDB2) N-malonyltransferase is structurally, functionally and phylogenetically distinct from its N-acetyltransferase (NAT) homologues.
- Author
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Karagianni EP, Kontomina E, Lowe ED, Athanasopoulos K, Papanikolaou G, Garefalaki V, Kotseli V, Zaliou S, Grimaud T, Arvaniti K, Tsatiri MA, Fakis G, Glenn AE, Roversi P, Abuhammad A, Ryan A, Sim RB, Sim E, and Boukouvala S
- Subjects
- Isoenzymes genetics, Phylogeny, Acetyl Coenzyme A, Acetyltransferases, Arylamine N-Acetyltransferase chemistry, Arylamine N-Acetyltransferase genetics, Fusarium genetics
- Abstract
Fusarium endophytes damage cereal crops and contaminate produce with mycotoxins. Those fungi overcome the main chemical defence of host via detoxification by a malonyl-CoA-dependent enzyme homologous to xenobiotic metabolizing arylamine N-acetyltransferase (NAT). In Fusarium verticillioides (teleomorph Gibberella moniliformis, GIBMO), this N-malonyltransferase activity is attributed to (GIBMO)NAT1, and the fungus has two additional isoenzymes, (GIBMO)NAT3 (N-acetyltransferase) and (GIBMO)NAT2 (unknown function). We present the crystallographic structure of (GIBMO)NAT1, also modelling other fungal NAT homologues. Monomeric (GIBMO)NAT1 is distinctive, with access to the catalytic core through two "tunnel-like" entries separated by a "bridge-like" helix. In the quaternary arrangement, (GIBMO)NAT1 monomers interact in pairs along an extensive interface whereby one entry of each monomer is covered by the N-terminus of the other monomer. Although monomeric (GIBMO)NAT1 apparently accommodates acetyl-CoA better than malonyl-CoA, dimerization changes the active site to allow malonyl-CoA to reach the catalytic triad (Cys110, His158 and Asp173) via the single uncovered entry, and anchor its terminal carboxyl-group via hydrogen bonds to Arg109, Asn157 and Thr261. Lacking a terminal carboxyl-group, acetyl-CoA cannot form such stabilizing interactions, while longer acyl-CoAs enter the active site but cannot reach catalytic Cys. Other NAT isoenzymes lack such structural features, with (GIBMO)NAT3 resembling bacterial NATs and (GIBMO)NAT2 adopting a structure intermediate between (GIBMO)NAT1 and (GIBMO)NAT3. Biochemical assays confirmed differential donor substrate preference of (GIBMO)NAT isoenzymes, with phylogenetic analysis demonstrating evolutionary separation. Given the role of (GIBMO)NAT1 in enhancing Fusarium pathogenicity, unravelling the structure and function of this enzyme may benefit research into more targeted strategies for pathogen control., (© 2022 Federation of European Biochemical Societies.)
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- 2023
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24. Investigation of the mycobacterial enzyme HsaD as a potential novel target for anti-tubercular agents using a fragment-based drug design approach.
- Author
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Ryan A, Polycarpou E, Lack NA, Evangelopoulos D, Sieg C, Halman A, Bhakta S, Eleftheriadou O, McHugh TD, Keany S, Lowe ED, Ballet R, Abuhammad A, Jacobs WR Jr, Ciulli A, and Sim E
- Subjects
- Antitubercular Agents chemical synthesis, Antitubercular Agents chemistry, Crystallography, X-Ray, Dose-Response Relationship, Drug, Enzyme Inhibitors chemical synthesis, Enzyme Inhibitors chemistry, Hydrolases deficiency, Hydrolases metabolism, Microbial Sensitivity Tests, Models, Molecular, Molecular Structure, Mycobacterium tuberculosis growth & development, Mycobacterium tuberculosis metabolism, Structure-Activity Relationship, Antitubercular Agents pharmacology, Drug Design, Enzyme Inhibitors pharmacology, Hydrolases antagonists & inhibitors, Mycobacterium tuberculosis drug effects, Mycobacterium tuberculosis enzymology
- Abstract
Background and Purpose: With the emergence of extensively drug-resistant tuberculosis, there is a need for new anti-tubercular drugs that work through novel mechanisms of action. The meta cleavage product hydrolase, HsaD, has been demonstrated to be critical for the survival of Mycobacterium tuberculosis in macrophages and is encoded in an operon involved in cholesterol catabolism, which is identical in M. tuberculosis and M. bovis BCG., Experimental Approach: We generated a mutant strain of M. bovis BCG with a deletion of hsaD and tested its growth on cholesterol. Using a fragment based approach, over 1000 compounds were screened by a combination of differential scanning fluorimetry, NMR spectroscopy and enzymatic assay with pure recombinant HsaD to identify potential inhibitors. We used enzymological and structural studies to investigate derivatives of the inhibitors identified and to test their effects on growth of M. bovis BCG and M. tuberculosis., Key Results: The hsaD deleted strain was unable to grow on cholesterol as sole carbon source but did grow on glucose. Of seven chemically distinct 'hits' from the library, two chemical classes of fragments were found to bind in the vicinity of the active site of HsaD by X-ray crystallography. The compounds also inhibited growth of M. tuberculosis on cholesterol. The most potent inhibitor of HsaD was also found to be the best inhibitor of mycobacterial growth on cholesterol-supplemented minimal medium., Conclusions and Implications: We propose that HsaD is a novel therapeutic target, which should be fully exploited in order to design and discover new anti-tubercular drugs., Linked Articles: This article is part of a themed section on Drug Metabolism and Antibiotic Resistance in Micro-organisms. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.14/issuetoc., (© 2017 The British Pharmacological Society.)
- Published
- 2017
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25. Identification of novel members of the bacterial azoreductase family in Pseudomonas aeruginosa.
- Author
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Crescente V, Holland SM, Kashyap S, Polycarpou E, Sim E, and Ryan A
- Subjects
- Anti-Bacterial Agents metabolism, Bacterial Proteins genetics, Coenzymes metabolism, Dinitrocresols metabolism, Models, Molecular, NAD(P)H Dehydrogenase (Quinone) metabolism, NADH, NADPH Oxidoreductases genetics, Niacinamide metabolism, Nitrofurazone metabolism, Nitroreductases, Oxidation-Reduction, Recombinant Proteins genetics, Substrate Specificity, Bacterial Proteins metabolism, NADH, NADPH Oxidoreductases metabolism, Pseudomonas aeruginosa metabolism, Recombinant Proteins metabolism
- Abstract
Azoreductases are a family of diverse enzymes found in many pathogenic bacteria as well as distant homologues being present in eukarya. In addition to having azoreductase activity, these enzymes are also suggested to have NAD(P)H quinone oxidoreductase (NQO) activity which leads to a proposed role in plant pathogenesis. Azoreductases have also been suggested to play a role in the mammalian pathogenesis of Pseudomonas aeruginosa. In view of the importance of P. aeruginosa as a pathogen, we therefore characterized recombinant enzymes following expression of a group of putative azoreductase genes from P. aeruginosa expressed in Escherichia coli. The enzymes include members of the arsenic-resistance protein H (ArsH), tryptophan repressor-binding protein A (WrbA), modulator of drug activity B (MdaB) and YieF families. The ArsH, MdaB and YieF family members all show azoreductase and NQO activities. In contrast, WrbA is the first enzyme to show NQO activity but does not reduce any of the 11 azo compounds tested under a wide range of conditions. These studies will allow further investigation of the possible role of these enzymes in the pathogenesis of P. aeruginosa., (© 2016 Authors; published by Portland Press Limited.)
- Published
- 2016
- Full Text
- View/download PDF
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