An approach to identifying novel substrates of bacterial arylamine N -acetyltransferases

Brooke, Edward W.; Davies, Stephen G.; Mulvaney, Andrew W.; Pompeo, Frédérique; Sim, Edith; Vickers, Richard

doi:10.1016/s0968-0896(02)00642-9

Cited by 84 publications

(87 citation statements)

References 31 publications

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“…The extreme preference for 2-aminophenols as acetyl acceptors is characteristic of SgNAT. This finding provides new insight into the substrate specificities of eukaryotic and bacterial NATs and should assist in identifying the endogenous roles of bacterial NATs (4).…”

mentioning

confidence: 83%

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ArylamineN-Acetyltransferase Responsible for Acetylation of 2-Aminophenols inStreptomyces griseus

2007

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An arylamine N-acetyltransferase (NAT) responsible for the N acetylation of exogenous 3-amino-4-hydroxybenzoic acid in Streptomyces griseus was identified and characterized. This enzyme was distinct from other eukaryotic and bacterial NATs in that it acetylated various 2-aminophenol derivatives more effectively than it acetylated 5-aminosalicylic acid, and thus it may be involved in the metabolism of xenobiotic compounds.

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mentioning

confidence: 83%

“…Of the substrates of bacterial NATs identified so far, 5-aminosalicylate (5-AS), a drug used in the treatment of inflammatory bowel diseases, is one of the most preferred substrates (6,24). Thus, bacterial NATs have the ability to detoxify xenobiotic compounds, and identification of bacterial NATs and their substrates still receives considerable attention (4).…”

mentioning

confidence: 99%

ArylamineN-Acetyltransferase Responsible for Acetylation of 2-Aminophenols inStreptomyces griseus

2007

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“…Enzyme Assays and Detection of Acetylated Aromatic Amines by HPLC-NAT activity was measured in the 5,5Ј-dithiobis-(2-nitrobenzoic acid) assay, as described previously (16). Recombinant enzymes and aromatic amine substrates (500 M final concentration) in assay buffer (25 mM Tris-HCl, pH 7.5) were incubated for 5 min at 37°C in a 96-well plate.…”

Section: Deletion Of Panat1 and Panat2 And Complementation Of The ⌬Pamentioning

confidence: 99%

An Acetyltransferase Conferring Tolerance to Toxic Aromatic Amine Chemicals

Martins

Rodrigues‐Lima

Dairou

et al. 2009

Journal of Biological Chemistry

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Aromatic amines (AA) are a major class of environmental pollutants that have been shown to have genotoxic and cytotoxic potentials toward most living organisms. Fungi are able to tolerate a diverse range of chemical compounds including certain AA and have long been used as models to understand general biological processes. Deciphering the mechanisms underlying this tolerance may improve our understanding of the adaptation of organisms to stressful environments and pave the way for novel pharmaceutical and/or biotechnological applications. We have identified and characterized two arylamine N-acetyltransferase (NAT) enzymes (PaNAT1 and PaNAT2) from the model fungus Podospora anserina that acetylate a wide range of AA. Targeted gene disruption experiments revealed that PaNAT2 was required for the growth and survival of the fungus in the presence of toxic AA. Functional studies using the knock-out strains and chemically acetylated AA indicated that tolerance of P. anserina to toxic AA was due to the N-acetylation of these chemicals by PaNAT2. Moreover, we provide proof-of-concept remediation experiments where P. anserina, through its PaNAT2 enzyme, is able to detoxify the highly toxic pesticide residue 3,4-dichloroaniline in experimentally contaminated soil samples. Overall, our data show that a single xenobiotic-metabolizing enzyme can mediate tolerance to a major class of pollutants in a eukaryotic species. These findings expand the understanding of the role of xenobiotic-metabolizing enzyme and in particular of NATs in the adaptation of organisms to their chemical environment and provide a basis for new systems for the bioremediation of contaminated soils.

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“…The measurement of NAT activity used pure recombinant mNat2 and the rate of hydrolysis of AcCoA in the presence of substrate was identified (Brooke et al 2003a). Inhibition of the hydrolysis of AcCoA was measured as described by Brooke et al (2003b).…”

Section: Activity Assaysmentioning

confidence: 99%

“…The rate of formation of coenzyme A (CoA) as a result of AcCoA hydrolysis was determined spectrophotometrically using the colorimetric agent 5,5 0 -dithio-bis (2-nitrobenzoic acid) (Ellman's reagent, DTNB) as previously described (Brooke et al 2003a), with the following modifications. The extent of reaction is measured by detecting the coloured 5-thio-2-nitrobenzoic acid, which is produced by the reaction of DTNB with free thiol CoA formed during the NAT reaction and has a maximum absorbance at 412 nm (Riddles et al 1983;Brooke et al 2003a). Samples of pure mNat2 (5 ng) were preincubated with pABA (500 mM final concentration) in assay buffer (20 mM Tris-HCl, pH 8.0) for 5 min at 378C in a 96-well plate (Corning).…”

Section: Activity Assaysmentioning

confidence: 99%

Prospective virtual screening with Ultrafast Shape Recognition: the identification of novel inhibitors of arylamine N -acetyltransferases

Ballester

Westwood

Laurieri

et al. 2009

J. R. Soc. Interface.

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There is currently a shortage of chemical molecules that can be used as bioactive probes to study molecular targets and potentially as starting points for drug discovery. One inexpensive way to address this problem is to use computational methods to screen a comprehensive database of small molecules to discover novel structures that could lead to alternative and better bioactive probes. Despite that pleasing logic the results have been somewhat mixed. Here we describe a virtual screening technique based on ligand -receptor shape complementarity, Ultrafast Shape Recognition (USR). USR is specifically applied to identify novel inhibitors of arylamine N-acetyltransferases by computationally screening almost 700 million molecular conformers in a time-and resource-efficient manner. A small number of the predicted active compounds were purchased and tested obtaining a confirmed hit rate of 40 per cent which is an outstanding result for a prospective virtual screening.

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An approach to identifying novel substrates of bacterial arylamine N -acetyltransferases

Cited by 84 publications

References 31 publications

ArylamineN-Acetyltransferase Responsible for Acetylation of 2-Aminophenols inStreptomyces griseus

ArylamineN-Acetyltransferase Responsible for Acetylation of 2-Aminophenols inStreptomyces griseus

An Acetyltransferase Conferring Tolerance to Toxic Aromatic Amine Chemicals

Prospective virtual screening with Ultrafast Shape Recognition: the identification of novel inhibitors of arylamine N -acetyltransferases

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