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Werf, A.J.M. van der

doi:10.1016/s0303-8467(75)80056-4

Cited by 3 publications

(6 citation statements)

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“…1. The requirement for energy in this reaction is consistent with the position of the equilibrium between 5-oxoproline and glutamate, which markedly favors the cyclic product (1)(2)(3). 5-Oxoprolinase activity has been found in a number of animal tissues and has been purified from kidney, a rich source of the enzyme.…”

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confidence: 55%

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Stimulation of hepatic glutathione formation by administration of L-2-oxothiazolidine-4-carboxylate, a 5-oxo-L-prolinase substrate.

Williamson¹,

Meister²

1981

Proc. Natl. Acad. Sci. U.S.A.

166

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5-Oxo-L-prolinase, the enzyme that catalyzes the conversion of 5-oxo-L-proline to L-glutamate coupled to the cleavage of ATP to ADP and Pi, also acts on L-2-oxothiazolidine-4-carboxylate (an analog of 5-oxoproline in which the 4-methylene moiety is replaced by sulfur) and ATP to yield cysteine and ADP. The enzyme, which exhibits an affinity for the analog similar to that for the natural substrate, is inhibited by the analog in vitro and in vivo. L-2-Oxothiazolidine-4-carboxylate thus serves as a ptent inhibitor of the y-glutamyl cycle at the step of 5-oxoprolinase. Administration of L-2-oxothiazolidine-4-carboxylate to mice that had been depleted of hepatic glutathione led to restoration of normal hepatic glutathione levels. Since L-2-oxothiazolidine-4-carboxylate is an excellent substrate of the enzyme, it may serve as an intracellular delivery system for cysteine and thus has potential as a therapeutic agent for conditions in which there is depletion of hepatic glutathione.5-Oxo-L-prolinase catalyzes the ATP-dependent hydrolysis of 5-oxo-L-proline according to the reaction given in Fig. 1. The requirement for energy in this reaction is consistent with the position of the equilibrium between 5-oxoproline and glutamate, which markedly favors the cyclic product (1-3). 5-Oxoprolinase activity has been found in a number of animal tissues and has been purified from kidney, a rich source of the enzyme. Earlier work in this laboratory established that 5-oxo-L-proline is a quantitatively significant metabolite of glutathione which is formed in the 'y-glutamyl cycle by the action of 'y-glutamyl cyclotransferase on y-glutamyl amino acids (4). Thus, the enzyme-catalyzed hydrolysis of 5-oxo-L-proline links the reactions involved in the utilization of glutathione (catalyzed by y-glutamyl transpeptidase, y-glutamyl cyclotransferase, and cysteinylglycinase) with those involved in its synthesis (catalyzed by y-glutamylcysteine synthetase and glutathione synthetase). In previous work (5) it was shown that L-2-imidazolidone4-carboxylate, a competitive inhibitor of 5-oxoprolinase, markedly decreases the metabolism of 5-oxoproline in vivo.Here we describe a new heterocyclic substrate of 5-oxoprolinase, L-2-oxothiazolidine-4-carboxylate, which is cleaved by the enzyme according to the scheme given in Fig. 1. In this pathway it is assumed that S-carboxy-cysteine, the initial product of hydrolysis, decarboxylates nonenzymatically. We have found that administration of L-2-oxothiazolidine-4-carboxylate to mice produces marked inhibition of the metabolism of 5-oxoproline but not that of glutamate. Administration of this compound also stimulates formation of glutathione in the liver.EXPERIMENTAL PROCEDURES Materials. The L and D isomers of 2-oxothiazolidine-4-carboxylate were synthesized by the method of Kaneko et al. (6) as modified (7). We are indebted to Sidney Weinhouse for a sample of the L isomer which was used in our initial studies. 5-Oxo-L-prolinase has been purified from rat kidney (3); the enzyme used in the present...

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confidence: 55%

“…The thiazolidine is substantially more active as an inhibitor than the most active previously known inhibitor, L-2-imidazolidone-4-carboxylate. The latter compound serves as a partial substrate for the enzyme; thus, it stimulates rapid hydrolysis of ATP but is not converted to an amino acid product (3,5,11).…”

Section: Discussionmentioning

confidence: 99%

Stimulation of hepatic glutathione formation by administration of L-2-oxothiazolidine-4-carboxylate, a 5-oxo-L-prolinase substrate.

Williamson¹,

Meister²

1981

Proc. Natl. Acad. Sci. U.S.A.

166

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“…'y-Glutamyl-cysteine synthetase (9) and 5-oxoprolinase (8) were purified from rat kidney. Rat kidney -y-glutamyl transpeptidase and sheep brain 7y-glutamylcyclotransferase were prepared by Suresh Tate and Vaira Wellner, respectively, of this laboratory.…”

mentioning

confidence: 99%

“…Previous studies showed that the enzyme is active toward 2-piperidone-6-carboxylate and the 3-and 4-hydroxy derivatives of 5-oxo-L-proline (8). L-2-Imidazolidone-4-carboxylate is a potent competitive inhibitor in vitro (8,23), and the effect of this compound has also been demonstrated in vlvo (6,23). It is of interest that several of the compounds studied here, which are not decyclized by the enzyme, nevertheless induce ATP cleavage (8,24,25).…”

mentioning

confidence: 99%

“…L-2-Imidazolidone-4-carboxylate is a potent competitive inhibitor in vitro (8,23), and the effect of this compound has also been demonstrated in vlvo (6,23). It is of interest that several of the compounds studied here, which are not decyclized by the enzyme, nevertheless induce ATP cleavage (8,24,25). Inhibition of L-y-glutamyl-L-a-aminobutyrylglycine synthesis was determined in 250-,ul reaction mixtures containing 100 mM imidazole-HCl (pH 7.2), 100 mM KCl, 10 mM ATP, 10 mM phosphoenolpyruvate, 20 mM MgCl2, 10 mM L-y-glutamyl-L-a-aminobutyrate, 10 (12,13,26).…”

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confidence: 99%

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Selective inhibition of gamma-glutamyl-cycle enzymes by substrate analogs.

Griffith¹,

Meister²

1977

Proc. Natl. Acad. Sci. U.S.A.

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Substrate analogs have been obtained that selectively inhibit the reactions of the -glutamyl cycle or that are susceptible to only limited metabolism by the cycle. Thus, glutathione synthesis may be inhibited and analogs of glutathione may be synthesized that do not participate in transpeptidation. Specific inhibitors of "y-glutamylcyclotransferase and 5-oxoprolinase have been obtained. The findings offer new approaches to the in vivo study of the cycle and also to the design of more specifically directed analogs of inhibitors such as methionine sulfoximine and 6-diazo-5-oxonorleucine.The enzymic reactions of the y-glutamyl cycle [which accounts for the synthesis and degradation of glutathione ( Fig. 1) (1)] include synthesis of the "y-glutamyl peptide bond, transfer of the y-glutamyl group to an acceptor, cyclization of the -y-glutamyl group to form 5-oxoproline, and energy-dependent decyclization of 5-oxoproline to glutamate. In an approach to further understanding of the function of the cycle, we have sought specific inhibitors of its individual reactions and also substrate analogs that would function in some but not all of the reactions. Such goals might be difficult to achieve because all of these reactions involve the y-carboxyl group of glutamate, and a given substrate analog might therefore be expected to interact with more than one enzyme. However, the several enzymic reaction pathways are clearly different and are presumably associated with differences in the enzyme-bound conformations of the glutamate carbon chain at the active sites of the several enzymes.In the present work, we have attempted to exploit such expected differences among the enzymes by manipulation of substrate structure.* We have found that suitable modification of the glutamyl moiety of the substrates yields the desired results: effective and specific inhibitors or nonmetabolizable analogs at each of the steps of the 'y-glutamyl cycle. The present findings thus offer several potentially new approaches to the study of the cycle in vvo and also provide information relevant to further investigations on the mechanisms of action of these metabolically related enzymes. The data reported here may be applied to the modification of potent (but relatively nonspecific) inhibitors such as methionine sulfoximine, azaserine, and 6-diazo-5-oxonorleucine so as to achieve one-enzyme specificity. The findings also make possible improved assay procedures for y-glutamyl-cysteine synthetase, glutathione synthetase, and y-glutamylcyclotransferase in which the substrates and products are not degraded by other enzymes present in unpurified tissue extracts. D-y-Glutamyl-L-a-aminobutyric acid, 'y-(y-methyl)glutamyl-L-a-aminobutyric acid, and fl-aminoglutaryl-L-a-aminobutyric acid were prepared via, the corresponding Nphthaloyl glutamic anhydride analogs as described (7). L-'y-glutamyl-La-aminobutyric acid, and f3-aminoglutaryl-L-a-aminobutyric acid were prepared enzymatically with -y-glutamyl-cysteine synthetase by using phosphoenolpyruvate and pyruv...

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Purification and Characterization of 5-Oxo-l-prolinase (l-Pyroglutamate Hydrolase) fromAlcaligenessp. F-137

Koyama

1988

Agricultural and Biological Chemistry

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Alcaligenes sp. F-137 has been purified to homogeneity as judged by acrylamide gel electrophoresis and ultracentrifugation. The molecular weight of the enzyme was about 106,000 and 123,000 by gel filtration and sedimentation equilibrium, respectively. Upon disc electrophoresis in 0.1 % sodium dodecyl sulfate the purified preparation migrates as a single band of molecular weight 126,000. The sedimentation coefficient (s~o. w) of the enzyme was 6.82 S by ultracentrifugation and its isoelectric point was pH 5.1 by isoelectric focusing. The enzyme catalyzed the hydrolysis of 5-oxo-L-proline to glutamate coupled with the hydrolysis of ATP to ADP and inorganic phosphate, stoichiometrically. K + and Mg2 + were required for the enzyme reaction. Michaelis constants of the enzyme were 0.07 mM for 5oxo-L-proline and 0.32mM for ATP. The enzyme was maximally active at pH 7.8 and 50°e. The enzyme was inhibited by p-chloromercuribenzoate of N-ethylmaleimide.

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Cited by 3 publications

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Stimulation of hepatic glutathione formation by administration of L-2-oxothiazolidine-4-carboxylate, a 5-oxo-L-prolinase substrate.

Stimulation of hepatic glutathione formation by administration of L-2-oxothiazolidine-4-carboxylate, a 5-oxo-L-prolinase substrate.

Selective inhibition of gamma-glutamyl-cycle enzymes by substrate analogs.

Purification and Characterization of 5-Oxo-l-prolinase (l-Pyroglutamate Hydrolase) fromAlcaligenessp. F-137

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