<i>N</i>
            -Acetylanthranilate Amidase from
            <i>Arthrobacter nitroguajacolicus</i>
            Rü61a, an α/β-Hydrolase-Fold Protein Active towards Aryl-Acylamides and -Esters, and Properties of Its Cysteine-Deficient Variant

Kolkenbrock, Stephan; Parschat, Katja; Beermann, Bernd; Hinz, Hans‐Jürgen; Fetzner, Susanne

doi:10.1128/jb.00325-07

Cited by 4 publications

(5 citation statements)

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“…DC-6 (Chen et al 2016 ), histone-like hydrolase PrpH (AEO21835) from Sphingomonas sp. Y57 (Zhang et al 2011 ), and α/β hydrolase Amq (YP_001210456) from Arthrobacter nitroguajacolicus Rü61a (Kolkenbrock et al 2007 ). However, the similarities between AmiH52 and these functional hydrolases obtained from the NCBI/Swiss-Prot database were quite low (less than 15%) after performing the multiple sequence alignment by CLUSTALW.…”

Section: Resultsmentioning

confidence: 99%

Characterization of a novel amidohydrolase with promiscuous esterase activity from a soil metagenomic library and its application in degradation of amide herbicides

Sun,

Chen,

Peng

et al. 2024

Environ Sci Pollut Res

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Amide herbicides have been extensively used worldwide and have received substantial attention due to their adverse environmental effects. Here, a novel amidohydrolase gene was identified from a soil metagenomic library using diethyl terephthalate (DET) as a screening substrate. The recombinant enzyme, AmiH52, was heterologously expressed in Escherichia coli and later purified and characterized, with the highest activity occurring at 40 ℃ and pH 8.0. AmiH52 was demonstrated to have both esterase and amidohydrolase activities, which exhibited highly specific activity for p-nitrophenyl butyrate (2669 U/mg) and degrading activity against several amide herbicides. In particular, it displayed the strongest activity against propanil, with a high degradation rate of 84% at 8 h. A GC–MS analysis revealed that propanil was transformed into 3,4-dichloroaniline (3,4-DCA) during this degradation. The molecular interactions and binding stability were then analyzed by molecular docking and molecular dynamics simulation, which revealed that several key amino acid residues, including Tyr164, Trp66, Ala59, Val283, Arg58, His33, His191, and His226, are involved in the specific interactions with propanil. This study provides a function-driven screening method for amide herbicide hydrolase from the metagenomic libraries and a promising propanil-degrading enzyme (AmiH52) for potential applications in environmental remediation.

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Section: Resultsmentioning

confidence: 99%

Characterization of a novel amidohydrolase with promiscuous esterase activity from a soil metagenomic library and its application in degradation of amide herbicides

Sun,

Chen,

Peng

et al. 2024

Environ Sci Pollut Res

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“…Assays with NATAc were carried out according to Kolkenbrock et al. [70]. The hydrolysis of NATAc was recorded for 30 min at 293 and 325 nm.…”

Section: Methodsmentioning

confidence: 99%

“…The hydrolysis of ATMA was recorded for 30 min at 290 nm (e o)NTMNPA = 1850 m )1 AEcm )1 ), and rate measurements were performed on the steady-state phase [26]. Assays with NATAc were carried out according to Kolkenbrock et al [70]. The hydrolysis of NATAc was recorded for 30 min at 293 and 325 nm.…”

Section: Acetanilidesmentioning

confidence: 99%

Kinetic analysis of effector modulation of butyrylcholinesterase‐catalysed hydrolysis of acetanilides and homologous esters

Masson¹,

Froment²,

Gillon³

et al. 2008

The FEBS Journal

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The effects of tyramine, serotonin and benzalkonium on the esterase and aryl acylamidase activities of wild-type human butyrylcholinesterase and its peripheral anionic site mutant, D70G, were investigated. The kinetic study was carried out under steady-state conditions with neutral and positively charged aryl acylamides [o-nitrophenylacetanilide, o-nitrotrifluorophenylacetanilide and m-(acetamido) N,N,N-trimethylanilinium] and homologous esters (o-nitrophenyl acetate and acetylthiocholine). Tyramine was an activator of hydrolysis for neutral substrates and an inhibitor of hydrolysis for positively charged substrates. The affinity of D70G for tyramine was lower than that of the wild-type enzyme. Tyramine activation of hydrolysis for neutral substrates by D70G was linear. Tyramine was found to be a pure competitive inhibitor of hydrolysis for positively charged substrates with both wild-type butyrylcholinesterase and D70G. Serotonin inhibited both esterase and aryl acylamidase activities for both positively charged and neutral substrates. Inhibition of wild-type butyrylcholinesterase was hyperbolic (i.e. partial) with neutral substrates and linear with positively charged substrates. Inhibition of D70G was linear with all substrates. A comparison of the effects of tyramine and serotonin on D70G versus the wild-type enzyme indicated that: (a) the peripheral anionic site is involved in the nonlinear activation and inhibition of the wild-type enzyme; and (b) in the presence of charged substrates, the ligand does not bind to the peripheral anionic site, so that ligand effects are linear, reflecting their sole interaction with the active site binding locus. Benzalkonium acted as an activator at low concentrations with neutral substrates. High concentrations of benzalkonium caused parabolic inhibition of the activity with neutral substrates for both wild-type butyrylcholinesterase and D70G, suggesting multiple binding sites. Benzalkonium caused linear, noncompetitive inhibition of the positively charged aryl acetanilide m-(acetamido) N,N,N-trimethylanilinium for D70G, and an unusual mixed-type inhibition ⁄ activation (a > b > 1) for wild-type butyrylcholinesterase with this substrate. No fundamental difference was observed between the effects of ligands on the butyrylcholinesterase-catalysed hydrolysis of esters and amides. Thus, Abbreviations

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“…In industry, one frequently tries to modify an enzyme in order to enhance its functionality in a certain way [1][2][3][4][5]. From an application point of view, one of the most interesting questions is how to modify an enzyme such that its activity is enhanced compared to wild type or such that a new kind of activity is introduced into the enzyme [6].…”

Section: Introductionmentioning

confidence: 99%

In silicoscreening of 393 mutants facilitates enzyme engineering of amidase activity in CalB

Hediger

Vico

Rannes

et al. 2013

PeerJ

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Our previously presented method for high throughput computational screening of mutant activity (Hediger et al., 2012) is benchmarked against experimentally measured amidase activity for 22 mutants of Candida antarctica lipase B (CalB). Using an appropriate cutoff criterion for the computed barriers, the qualitative activity of 15 out of 22 mutants is correctly predicted. The method identifies four of the six most active mutants with ≥3-fold wild type activity and seven out of the eight least active mutants with ≤0.5-fold wild type activity. The method is further used to screen all sterically possible (386) double-, triple- and quadruple-mutants constructed from the most active single mutants. Based on the benchmark test at least 20 new promising mutants are identified.

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N -Acetylanthranilate Amidase from Arthrobacter nitroguajacolicus Rü61a, an α/β-Hydrolase-Fold Protein Active towards Aryl-Acylamides and -Esters, and Properties of Its Cysteine-Deficient Variant

Cited by 4 publications

References 0 publications

Characterization of a novel amidohydrolase with promiscuous esterase activity from a soil metagenomic library and its application in degradation of amide herbicides

Characterization of a novel amidohydrolase with promiscuous esterase activity from a soil metagenomic library and its application in degradation of amide herbicides

Kinetic analysis of effector modulation of butyrylcholinesterase‐catalysed hydrolysis of acetanilides and homologous esters

In silicoscreening of 393 mutants facilitates enzyme engineering of amidase activity in CalB

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