16 results on '"Sim, Edith"'
Search Results
2. Probing the architecture of the Mycobacterium marinum arylamine N-acetyltransferase active site
- Author
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Abuhammad, Areej M., Lowe, Edward D., Fullam, Elizabeth, Noble, Martin, Garman, Elspeth F., and Sim, Edith
- Published
- 2010
- Full Text
- View/download PDF
3. 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, 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, William R, Ciulli, Alessio, Sim, Edith, and Jacobs, William R Jr
- Subjects
DRUG resistance ,TUBERCULOSIS ,MACROPHAGES ,MYCOBACTERIUM ,OPERONS ,ANIMAL experimentation ,ANTITUBERCULAR agents ,BIOCHEMISTRY ,CHEMISTRY ,DOSE-effect relationship in pharmacology ,ENZYME inhibitors ,HYDROLASES ,MATHEMATICAL models ,PHENOMENOLOGY ,MICROBIAL sensitivity tests ,MOLECULAR structure ,MYCOBACTERIUM tuberculosis ,RESEARCH funding ,DRUG development ,THEORY ,CHEMICAL inhibitors ,PHARMACODYNAMICS - 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. [ABSTRACT FROM AUTHOR]- Published
- 2017
- Full Text
- View/download PDF
4. Exploration of Piperidinols as Potential Antitubercular Agents.
- Author
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Abuhammad, Areej, Fullam, Elizabeth, Bhakta, Sanjib, Russell, Angela J., Morris, Garrett M., Finn, Paul W., and Sim, Edith
- Subjects
ANTITUBERCULAR agents ,TUBERCULOSIS treatment ,ARYLAMINE N-acetyltransferase ,MYCOBACTERIUM marinum ,AMINO acids - Abstract
Novel drugs to treat tuberculosis are required and the identification of potential targets is important. Piperidinols have been identified as potential antimycobacterial agents (MIC < 5 μg/mL), which also inhibit mycobacterial arylamine N-acetyltransferase (NAT), an enzyme essential for mycobacterial survival inside macrophages. The NAT inhibition involves a prodrug-like mechanism in which activation leads to the formation of bioactive phenyl vinyl ketone (PVK). The PVK fragment selectively forms an adduct with the cysteine residue in the active site. Time dependent inhibition of the NAT enzyme from Mycobacterium marinum (M. marinum) demonstrates a covalent binding mechanism for all inhibitory piperidinol analogues. The structure activity relationship highlights the importance of halide substitution on the piperidinol benzene ring. The structures of the NAT enzymes from M. marinum and M. tuberculosis, although 74% identical, have different residues in their active site clefts and allow the effects of amino acid substitutions to be assessed in understanding inhibitory potency. In addition, we have used the piperidinol 3-dimensional shape and electrostatic properties to identify two additional distinct chemical scaffolds as inhibitors of NAT. While one of the scaffolds has anti-tubercular activity, both inhibit NAT but through a non-covalent mechanism. [ABSTRACT FROM AUTHOR]
- Published
- 2014
- Full Text
- View/download PDF
5. Pathogenic mycobacteria achieve cellular persistence by inhibiting the Niemann-Pick Type C disease cellular pathway
- Author
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Fineran, Paul, Lloyd-Evans, Emyr, Lack, Nathan A., Platt, Nick, Davis, Lianne C., Morgan, Anthony J., Höglinger, Doris, Tatituri, Raju Venkata V., Clark, Simon, Williams, Ian M., Tynan, Patricia, Al Eisa, Nada, Nazarova, Evgeniya, Williams, Ann, Galione, Antony, Ory, Daniel S., Besra, Gurdyal S., Russell, David G., Brenner, Michael B., Sim, Edith, and Platt, Frances M.
- Subjects
Tuberculosis ,Niemann-Pick Disease Type C ,Lysosomal Storage Diseases ,Lysosomal Calcium - 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 persistent 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 persistent 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 persistent 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.
- Published
- 2016
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6. Structure of arylamine N-acetyltransferase from Mycobacterium tuberculosis determined by cross-seeding with the homologous protein from M. marinum: triumph over adversity.
- Author
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Abuhammad, Areej, Lowe, Edward D., McDonough, Michael A., Shaw Stewart, Patrick D., Kolek, Stefan A., Sim, Edith, and Garman, Elspeth F.
- Subjects
ARYLAMINE N-acetyltransferase ,MYCOBACTERIUM tuberculosis ,MYCOBACTERIUM marinum ,MACROPHAGES ,PHARMACEUTICAL chemistry - Abstract
Arylamine N-acetyltransferase from Mycobacterium tuberculosis (TBNAT) plays an important role in the intracellular survival of the microorganism inside macrophages. Medicinal chemistry efforts to optimize inhibitors of the TBNAT enzyme have been hampered by the lack of a three-dimensional structure of the enzyme. In this paper, the first structure of TBNAT, determined using a lone crystal produced using cross-seeding with the homologous protein from M. marinum, is reported. Despite the similarity between the two enzymes (74% sequence identity), they show distinct physical and biochemical characteristics. The structure elegantly reveals the characteristic features of the protein surface as well as details of the active site of TBNAT relevant to drug-discovery efforts. The crystallographic analysis of the diffraction data presented many challenges, since the crystal was twinned and the habit possessed pseudo-translational symmetry. [ABSTRACT FROM AUTHOR]
- Published
- 2013
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- View/download PDF
7. Characterization of an oxidoreductase from the arylamine N-acetyltransferase operon in Mycobacterium smegmatis.
- Author
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Evangelopoulos, Dimitrios, Cronin, Nora, Daviter, Tina, Sim, Edith, Keep, Nicholas H., and Bhakta, Sanjib
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OXIDOREDUCTASES ,ACETYLTRANSFERASES ,MYCOBACTERIA ,GENETIC regulation ,OPERONS ,MYCOBACTERIAL diseases ,TUBERCULOSIS ,PROTEIN structure - Abstract
Mycobacterium tuberculosis, the most successful bacterial pathogen, causes tuberculosis, a disease that still causes more than 2 million deaths per year. Arylamine N-acetyltransferase is an enzyme that is conserved in most Mycobacterium spp. The nat gene belongs to an operon that is important for the intracellular survival of M. tuberculosis within macrophages. The nat operon in Mycobacterium smegmatis and other fast-growing mycobacterial species has a unique organization containing genes with uncharacterized function. Here, we describe the biochemical, biophysical and structural characterization of the MSMEG_0308 gene product (MS0308) of the M. smegmatis nat operon. While characterizing the function of MS0308, we validated the oxidoreductase property; however, we found that the enzyme was not utilizing dihydrofolate as its substrate, hence we first report that MS0308 is not a dihydrofolate reductase, as annotated in the genome. The structure of this oxidoreductase was solved at 2.0 Å in complex with the cofactor NADPH and has revealed the hydrophobic pocket where the endogenous substrate binds. Database The structural information for the MS0308 oxidoreductase has been deposited to the Protein Data Bank under accession number . Structured digital abstract [ABSTRACT FROM AUTHOR]
- Published
- 2011
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8. Arylamine N-acetyltransferases: Structural and functional implications of polymorphisms
- Author
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Sim, Edith, Lack, Nathan, Wang, Chan-Ju, Long, Hilary, Westwood, Isaac, Fullam, Elizabeth, and Kawamura, Akane
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ACETYLTRANSFERASES , *GENETIC polymorphisms , *ACETYLATION , *ACETYLCOENZYME A , *PHARMACOGENOMICS , *CATALYSIS , *EXONS (Genetics) , *ESTROGEN receptors , *ISONIAZID - Abstract
Abstract: Arylamine N-acetyltransferases (NATs) catalyse the N-acetylation of arylamines, arylhydroxylamines and arylhydrazines with the acetyl group being transferred from acetylCoenzyme A. As a result of many recent advances in NAT research there have been many recent reviews and the present paper gives a flavour of the excitement in the field. The NATs, which are cytosolic, were early examples of pharmacogenetic variation. Polymorphism in isoniazid inactivation resulting in slow acetylation was subsequently found to be due to SNPs in the gene encoding the human isoenzyme NAT2. There are two polymorphic genes (NAT1 and NAT2) encoded with a third pseudogene (NATP) at human 8p21.3. The gene structure of NAT1 and NAT2, with a single (NAT2) or multiple (NAT1) distant non-coding exons showing tissue specific splicing, opens possibilities for effects of polymorphisms outside the single coding exon. In humans, the substrate specificities of NAT1 and NAT2 are overlapping but distinct. The NAT2 isoenzyme, predominantly in liver and gut, acetylates sulphamethazine and arylhydrazine compounds. Slow acetylators are at increased risk of toxicity, e.g. isoniazid induced neurotoxicity and hydralazine-induced lupus. The human NAT1 isoenzyme is also polymorphic. It is expressed in many tissues, particularly in oestrogen receptor positive breast cancers. Human NAT1 has an endogenous role in acetylation of a folate catabolite with in vivo evidence from transgenic mice lacking the equivalent gene. For nomenclature see http://louisville.edu/medschool/pharmacology/NAT.html, the website maintained by David Hein. NAT homologues have been identified by bioinformatics analyses in zebrafish and these sequences are described, although the proteins have not yet been characterized. The first NAT crystallographic structure from Salmonella typhimurium identified the mechanism of acetyl transfer via a catalytic triad of Cys, His and Asp residues each essential for activity in all NATs. NATs from mycobacteria aided in identifying the substrate binding site and the acetylCoA binding pocket. Studies on the eukaryotic enzymes by NMR and crystallography have facilitated understanding substrate specificities of human NAT1 (5-aminosalicylate and p-aminobenzoic acid) and human NAT2 (sulphamethazine). The effect of “slow acetylator” SNPs in the coding region predominantly act through creating unstable protein that aggregates intracellularly prior to ubiquitination and degradation. [Copyright &y& Elsevier]
- Published
- 2008
- Full Text
- View/download PDF
9. Binding of the anti-tubercular drug isoniazid to the arylamine N-acetyltransferase protein from Mycobacterium smegmatis.
- Author
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Sandy, James, Holton, Simon, Fullam, Elizabeth, Sim, Edith, and Noble, Martin
- Abstract
Isoniazid is a frontline drug used in the treatment of tuberculosis (TB). Isoniazid is a prodrug, requiring activation in the mycobacterial cell by the catalase/peroxidase activity of the katG gene product. TB kills two million people every year and the situation is getting worse due to the increase in prevalence of HIV/AIDS and emergence of multidrug-resistant strains of TB. Arylamine N-acetyltransferase (NAT) is a drug-metabolizing enzyme (E.C. 2.1.3.5). NAT can acetylate isoniazid, transferring an acetyl group from acetyl coenzyme A onto the terminal nitrogen of the drug, which in its N-acetylated form is therapeutically inactive. The bacterium responsible for TB, Mycobacterium tuberculosis, contains and expresses the gene encoding the NAT protein. Isoniazid binds to the NAT protein from Salmonella typhimurium and we report here the mode of binding of isoniazid in the NAT enzyme from Mycobacterium smegmatis, closely related to the M. tuberculosis and S. typhimurium NAT enzymes. The mode of binding of isoniazid to M. smegmatis NAT has been determined using data collected from two distinct crystal forms. We can say with confidence that the observed mode of binding of isoniazid is not an artifact of the crystallization conditions used. The NAT enzyme is active in mycobacterial cells and we propose that isoniazid binds to the NAT enzyme in these cells. NAT activity in M. tuberculosis is likely therefore to modulate the degree of activation of isoniazid by other enzymes within the mycobacterial cell. The structure of NAT with isoniazid bound will facilitate rational drug design for anti-tubercular therapy. [ABSTRACT FROM AUTHOR]
- Published
- 2005
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10. Stereoselective synthesis of β-arabino glycosyl sulfones as potential inhibitors of mycobacterial cell wall biosynthesis
- Author
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Ayers, Benjamin, Long, Hilary, Sim, Edith, Smellie, Iain A., Wilkinson, Brendan L., and Fairbanks, Antony J.
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INHIBITORS of bacterial cell wall synthesis , *SULFONES , *MONOSACCHARIDES , *MYCOBACTERIUM bovis , *BCG vaccines , *GLYCOSYLATION - Abstract
Abstract: A series of β-arabino glycosyl sulfones with varying alkyl chain lengths were synthesised in a stereoselective fashion as putative mimics of decaprenolphosphoarabinose (DPA), and as potential inhibitors of mycobacterial cell wall biosynthesis. Biological testing against Mycobacterium bovis BCG revealed low to moderate anti-mycobacterial activity with marked dependence on alkyl chain length, which was maximal for a C-12 chain. [Copyright &y& Elsevier]
- Published
- 2009
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11. Synthesis of Arabino glycosyl triazoles as potential inhibitors of mycobacterial cell wall biosynthesis
- Author
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Wilkinson, Brendan L., Long, Hilary, Sim, Edith, and Fairbanks, Antony J.
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TRIAZOLES , *MYCOBACTERIUM , *BIOSYNTHESIS , *BACTERIAL cell walls , *MYCOBACTERIA , *TUBERCULOSIS , *ORGANIC synthesis - Abstract
Abstract: A series of arabino glycosyl triazoles with varying hydrophobic groups were synthesised as putative mimics of decaprenolphosphoarabinose (DPA) as potential inhibitors of mycobacterial cell wall biosynthesis. Biological testing against Mycobacterium bovis BCG revealed low to moderate anti-mycobacterial activity, with strong dependence on the identity of the hydrophobic side chain. [Copyright &y& Elsevier]
- Published
- 2008
- Full Text
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12. Protein-ligand interactions of arylamine N-acetyltransferase from Mycobacterium smegmatis
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Brooke, Edward W. and Sim, Edith
- Subjects
616.995061 ,Tuberculosis ,Acetyltransferases ,Mycobacterium tuberculosis - Abstract
Tuberculosis is the world's largest cause of death from an infectious agent. Treatment is by an extended period of combination chemotherapy. Drug resistance is an increasing problem in tuberculosis therapy, particularly to the frontline anti-tubercular drug isoniazid (INH). Recombinant arylamine N-acetyltransferase (NAT) of Mycobacterium tuberculosis N-acetylates INH using the cofactor Acetyl Coenzyme A. NAT from M. tuberculosis is a polymorphic enzyme and also acetylates INH in vivo. Acetylated INH is inactive therapeutically against M. tuberculosis both in vivo and in vitro. The acetylation of isoniazid in the mycobacterial cell may compete with the activation of INH by the catalase-peroxidase, katG, and hence contribute to INH resistance in clinical isolates. Inhibition of NAT in M. tuberculosis may thus increase the efficacy of INH therapy. A novel assay based around the detection of free Coenzyme A released during the acetylation reaction was used to determine the substrate specificity of recombinant NAT from the related Mycobacterium M. smegmatis (MSNAT). A relationship was observed between the lipophilicity of simple arylamine substrates and the rate of acetylation by MSNAT. Several MSNAT substrates possess antibacterial activity. The assay could also be used to screen compound libraries for MSNAT inhibitors. Synthesis of seventeen thiazolidinedione sultams in collaboration with Dr.Vickers (Dyson Perrins), identified as weak inhibitors of MSNAT, gave a minimum competitive inhibitory constant of 14μM. Screening a library of 5,074 drug-like compounds for inhibition of MSNAT identified thirteen compounds with semi-maximal inhibition constants (IC
50 ) of below 10μM. Based on this, fifteen maleimides were synthesised and were irreversible inhibitors of MSNAT with submicromolar potency. Similarly, ninety-six aminothiazoles were synthesised by Dr. Vickers and were uncompetitive inhibitors of MSNAT with a minimum IC50 of 1.5μM. The most potent aminothiazole showed no effect on the growth of M. smegmatis or M. bovis BCG or the sensitivity of the bacteria to isoniazid. However the aminothiazoles were shown not to penetrate the cells.- Published
- 2003
13. Analysis of β-amino alcohols as inhibitors of the potential anti-tubercular target N-acetyltransferase
- Author
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Fullam, Elizabeth, Abuhammad, Areej, Wilson, David L., Anderton, Matthew C., Davies, Steve G., Russell, Angela J., and Sim, Edith
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PHARMACEUTICAL research , *AMINO alcohols , *ENZYME inhibitors , *BIOSYNTHESIS , *DRUG synergism , *ANTITUBERCULAR agents , *TARGETED drug delivery , *ACETYLTRANSFERASES , *MYCOBACTERIUM tuberculosis - Abstract
Abstract: The synthesis and inhibitory potencies of a novel series of β-amino alcohols, based on the hit-compound 3-[3′-(4″-cyclopent-2‴-en-1‴-ylphenoxy)-2′-hydroxypropyl]-5,5 dimethylimidazolidine-2,4-dione as specific inhibitors of mycobacterial N-acetyltransferase (NAT) enzymes are reported. Effects of synthesised compounds on growth of Mycobacterium tuberculosis have been determined. [Copyright &y& Elsevier]
- Published
- 2011
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14. Inhibition of mycobacterial arylamine N-acetyltransferase contributes to anti-mycobacterial activity of Warburgia salutaris
- Author
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Madikane, Vukani Eliya, Bhakta, Sanjib, Russell, Angela J., Campbell, William E., Claridge, Timothy D.W., Elisha, B. Gay, Davies, Stephen G., Smith, Peter, and Sim, Edith
- Subjects
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ACETYLTRANSFERASES , *AMINES , *ANTIBACTERIAL agents , *MYCOBACTERIAL disease treatment - Abstract
Abstract: In this study, we show that extracts and a purified compound of Warburgia salutaris exhibit anti-mycobacterial activity against Mycobacterium tuberculosis H37Rv and Mycobacterium bovis BCG Pasteur. The extracts did not inhibit growth of Escherichia coli and were not toxic to cultured mammalian macrophage cells at the concentrations at which anti-mycobacterial activity was observed. The extract and pure compound inhibited pure recombinant arylamine N-acetyltransferase (NAT), an enzyme involved in mycobacterial cell wall lipid synthesis. Moreover, neither extract nor pure compound inhibited growth of a strain of M. bovis BCG in which nat has been deleted suggesting that NAT may indeed be a target within the mycobacterial cell. The purified compound is a novel drimane sesquiterpenoid lactone, 11α-hydroxycinnamosmolide. These studies show that W. salutaris is a useful source of anti-tubercular compounds for further analysis and supports the hypothesis of a link between NAT inhibition and anti-mycobacterial activity. [Copyright &y& Elsevier]
- Published
- 2007
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15. The Structure of Arylamine N-acetyltransferase from Mycobacterium smegmatis—An Enzyme which Inactivates the Anti-tubercular Drug, Isoniazid
- Author
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Sandy, James, Mushtaq, Adeel, Kawamura, Akane, Sinclair, John, Sim, Edith, and Noble, Martin
- Subjects
- *
ACETYLTRANSFERASES , *TUBERCULOSIS - Abstract
Arylamine N-acetyltransferases which acetylate and inactivate isoniazid, an anti-tubercular drug, are found in mycobacteria including Mycobacterium smegmatis and Mycobacterium tuberculosis. We have solved the structure of arylamine N-acetyltransferase from M. smegmatis at a resolution of 1.7 A˚ as a model for the highly homologous NAT from M. tuberculosis. The fold closely resembles that of NAT from Salmonella typhimurium, with a common catalytic triad and domain structure that is similar to certain cysteine proteases. The detailed geometry of the catalytic triad is typical of enzymes which use primary alcohols or thiols as activated nucleophiles. Thermal mobility and structural variations identify parts of NAT which might undergo conformational changes during catalysis. Sequence conservation among eubacterial NATs is restricted to structural residues of the protein core, as well as the active site and a hinge that connects the first two domains of the NAT structure. The structure of M. smegmatis NAT provides a template for modelling the structure of the M. tuberculosis enzyme and for structure-based ligand design as an approach to designing anti-TB drugs. [Copyright &y& Elsevier]
- Published
- 2002
- Full Text
- View/download PDF
16. 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
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