Purification of Glutamine Transaminase K/Cysteine Conjugate β-Lyase from Rat Renal Cytosol Based on Hydrophobic Interaction HPLC and Gel Permeation FPLC

Yamauchi, Aiko; Stijntjes, G.J.; Commandeur, Jan N. M.; Vermeulen, Nico

doi:10.1006/prep.1993.1073

Cited by 22 publications

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“…As was previously noted for the rat enzyme [6,9], our data show that rhGTK catalyzes β-elimination more effectively than transamination when TFEC and DCVC are substrates (compare Table 1 and 3). Yamauchi et al…”

Section: Nih-pa Author Manuscriptsupporting

confidence: 86%

“…Se-Methyl-L-selenocysteine is >100 times more effective as an aminotransferase substrate than is L-selenomethionine despite the difference of only one methylene group. Thus, despite the ability of rhGTK to bind large amino acids there are subtle constraints in the geometry of amino acids that can bind most effectively at the active site.As was previously noted for the rat enzyme [6,9], our data show that rhGTK catalyzes β-elimination more effectively than transamination when TFEC and DCVC are substrates (compare Table 1 and 3). Yamauchi et al…”

supporting

confidence: 86%

“…Important biological roles include salvage of the α-keto acid analogues of several essential amino acids and closure of the last step of the methionine salvage pathway (reviewed in [5]). GTK also possesses β-lyase activity toward many cysteine S-conjugates that possess a strong electron-withdrawing group attached at the sulfur [5][6][8][9][10] (1)Rat kidney GTK catalyzes β-elimination reactions with S-(1,2-dichlorovinyl)-L-cysteine (DCVC; the cysteine S-conjugate of trichloroethylene), S-(1,1,2,2-tetrafluoroethyl)-L-cysteine (TFEC; the cysteine S-conjugate of tetrafluoroethylene) and many other cysteine S-conjugates derived from halogenated alkenes [5][6][8][9][10]. However, when these halogenated cysteine Sconjugates are substrates, transamination competes with the β-lyase reaction.…”

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confidence: 99%

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confidence: 99%

“…Rat kidney GTK catalyzes β-elimination reactions with S-(1,2-dichlorovinyl)-L-cysteine (DCVC; the cysteine S-conjugate of trichloroethylene), S-(1,1,2,2-tetrafluoroethyl)-L-cysteine (TFEC; the cysteine S-conjugate of tetrafluoroethylene) and many other cysteine S-conjugates derived from halogenated alkenes [5][6][8][9][10]. However, when these halogenated cysteine Sconjugates are substrates, transamination competes with the β-lyase reaction.…”

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Substrate specificity of human glutamine transaminase K as an aminotransferase and as a cysteine S-conjugate β-lyase

Cooper

Pinto

Krasnikov

et al. 2008

Archives of Biochemistry and Biophysics

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Rat kidney glutamine transaminase K (GTK) exhibits broad specificity both as an aminotransferase and as a cysteine S-conjugate β-lyase. The β-lyase reaction products are pyruvate, ammonium and a sulfhydryl-containing fragment. We show here that recombinant human GTK (rhGTK) also exhibits broad specificity both as an aminotransferase and as a cysteine S-conjugate β-lyase. S-(1,1,2,2-Tetrafluoroethyl)-L-cysteine is an excellent aminotransferase and β-lyase substrate of rhGTK. Moderate aminotransferase and β-lyase activities occur with the chemopreventive agent Se-methyl-L-selenocysteine. L-3-(2-Naphthyl)alanine, L-3-(1-naphthyl)alanine, 5-S-L-cysteinyldopamine and 5-S-L-cysteinyl-L-DOPA are measurable aminotransferase substrates, indicating that the active site can accommodate large aromatic amino acids. The α-keto acids generated by transamination/Lamino acid oxidase activity of the two catechol cysteine S-conjugates are unstable. A slow rhGTKcatalyzed β-elimination reaction, as measured by pyruvate formation, was demonstrated with 5-S-L-cysteinyldopamine, but not with 5-S-L-cysteinyl-L-DOPA. The importance of transamination, oxidation and β-elimination reactions involving 5-S-L-cysteinyldopamine, 5-S-L-cysteinyl-L-DOPA and Se-methyl-L-selenocysteine in human tissues and their biological relevance are discussed. KeywordsCysteine S-conjugate β-lyase; 5-S-L-cysteinyl-L-DOPA; 5-S-L-cysteinyldopamine; glutamine transaminase K; Se-methyl-L-selenocysteine Glutamine transaminase K (GTK) 1 purified from rat and bovine tissues accepts a large number of neutral, aromatic and sulfur/selenium-containing substrates [1][2][3][4][5][6]. GTK catalyzes *Corresponding author: Department of Biochemistry and Molecular Biology, New York Medical College, Valhalla, NY 10595, USA. Fax: +1 914 594 4058, E-mail address: arthur_cooper@nymc.edu. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. 1 Abbreviations used: BTC, benzothiazolyl-L-cysteine; DCVC, S-(1,2-dichlorovinyl)-L-cysteine; DOPA, dihydroxyphenylalanine; DTNB, 5,5′-dithiobis(2-nitrobenzoic acid); DTT, dithiothreitol; GSH, glutathione; KAT, kynurenine aminotransferase; KGDHC, α-ketoglutarate dehydrogenase complex; KGM, α-ketoglutaramate; KMB, α-keto-γ-methiolbutyrate; MBTH, 3-methyl-2-benzothiazolinone hydrazone; MDH, malate dehydrogenase; MPA, metaphosphoric acid; PDHC, pyruvate dehydrogenase complex; PLP, pyridoxal 5′-phosphate; rhGTK, recombinant human glutamine transaminase K; TFEC, S- (1,1,2,2-tetrafluoroethyl) NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript transamination of kynurenine to kynurenate, ...

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Section: Nih-pa Author Manuscriptsupporting

confidence: 86%

supporting

confidence: 86%

mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Substrate specificity of human glutamine transaminase K as an aminotransferase and as a cysteine S-conjugate β-lyase

Cooper

Pinto

Krasnikov

et al. 2008

Archives of Biochemistry and Biophysics

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Measurement of Cysteine S‐Conjugate β‐Lyase Activity

et al. 2010

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Cysteine S-conjugate β-lyases are pyridoxal 5′-phosphate (PLP)-containing enzymes that catalyze the conversion of cysteine S-conjugates [RSCH 2 CH(NH 3 + )CO 2 − ] and selenium Se-conjugates[RSeCH 2 CH(NH 3 + )CO 2 − ] that contain a leaving group in the β position to pyruvate, ammonium and a sulfur-containing fragment (RSH) or selenium-containing fragment (RSeH), respectively. At least ten PLP enzymes catalyze β-elimination reactions with such cysteine S-conjugates. All are enzymes involved in amino acid metabolism that do not normally catalyze a β-lyase reaction, but catalyze a non-physiological β-lyase side reaction that depends on the electron-withdrawing properties of the -SR or -SeR moiety. In the case of the cysteine S-conjugates, if the eliminated RSH is stable the compound may be S-thiomethylated and excreted (thiomethyl shunt) or Sglucuronidated and harmlessly excreted [the possibility that RSeH compounds may be similarly metabolized has not been extensively studied]. If, however, RSH is chemically reactive the cysteine S-conjugate may be toxic as a result of the β-lyase reaction. The cysteine S-conjugate β-lyase pathway is of particular interest to toxicologists because it is involved in the bioactivation (toxification) of halogenated alkenes and certain drugs. Keywords Ammonium; cysteine S-conjugates; cysteine S-conjugate β-lyases; S-(1,2-dichlorovinyl)-Lcysteine; S-(1,1,2,2-tetrafluoroethyl)-L-cysteine; pyruvateMany potentially toxic electrophiles, whether of endogenous origin (e.g. α,β-unsaturated aldehydes derived from the oxidation of lipids) or of exogenous origin (e.g. some drugs), are detoxified through the mercapturate pathway (Figure 1). In addition, some non-electrophilic toxins may be converted to electrophiles that are then detoxified through the mercapturate pathway. The electrophile is sequentially converted to the glutathione S-conjugate, cysteinylglycine S-conjugate, cysteinyl S-conjugate and N-acetyl cysteine S-conjugate (mercapturate) by the action of glutathione S-transferases, γ-glutamyltransferase (γ-glutamyltranspeptidase), dipeptidases and N-acetyl transferases, respectively. The mercapturate is generally more hydrophilic and water soluble than is the starting electrophile and is readily excreted in the urine or bile.If the cysteine S-conjugate contains a good leaving group (nucleofuge) in the β position the metabolism of the electrophile may be diverted irreversibly away from the mercapturate by the action of cysteine S-conjugate β-lyases. These PLP-containing enzymes catalyze the conversion of the cysteine S-conjugate [RSCH 2 CH(NH 3 + )CO 2 − ] to RSH and aminoacrylate

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Glutathione‐Dependent Bioactivation

Lash

2007

CP Toxicology

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The classical view of the glutathione (GSH) conjugation pathway involves GSH S-transferase (GST)-dependent formation of thioether conjugates between GSH and an electrophilic substrate, processing to yield the corresponding cysteine S-conjugate, which is then converted to an N-acetylcysteine conjugate (or mercapturate). Mercapturates of most GST substrates are rendered more polar and thus readily excreted in urine. In contrast, there is a growing number of GST substrates that, rather than being detoxified, are bioactivated. These substrates include several halogenated solvents, many of which are nephrotoxic because of the tissue distribution of GSH conjugation pathway enzymes and membrane transporters, and prodrugs of certain chemotherapeutic agents. Although the initiating steps are the same regardless of whether the substrate is detoxified or bioactivated, the cysteine conjugate functions as a branch point. Bioactivated cysteine S-conjugates are metabolized in the kidneys by either cysteine conjugate β-lyase or flavin-containing monooxygenase to produce a reactive intermediate.

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Purification of Glutamine Transaminase K/Cysteine Conjugate β-Lyase from Rat Renal Cytosol Based on Hydrophobic Interaction HPLC and Gel Permeation FPLC

Cited by 22 publications

References 0 publications

Substrate specificity of human glutamine transaminase K as an aminotransferase and as a cysteine S-conjugate β-lyase

Substrate specificity of human glutamine transaminase K as an aminotransferase and as a cysteine S-conjugate β-lyase

Measurement of Cysteine S‐Conjugate β‐Lyase Activity

Glutathione‐Dependent Bioactivation

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