Kinetic studies on rat liver microsomal glutathione transferase: consequences of activation

Andersson, Claes; Piemonte, Fiorella; Mosialou, Erifili; Weinander, Rolf; Sun, Tie Hua; Lundqvist, Gerd; Adang, Anton E.P.; Morgenstern, Ralf

doi:10.1016/0167-4838(94)00239-d

Cited by 23 publications

(28 citation statements)

References 26 publications

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“…Our data demonstrate that R70 is important to the transferase mechanism while not critically involved in substrate binding. WT EXP1 reaction kinetics were in good agreement with other eukaryotic MGSTs (Andersson et al, 1995). …”

Section: Resultssupporting

confidence: 69%

Supergenomic Network Compression and the Discovery of EXP1 as a Glutathione Transferase Inhibited by Artesunate

Lisewski

Quiros

et al. 2014

Cell

113

128

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Summary A central problem in biology is to identify gene function. One approach is to infer function in large supergenomic networks of interactions and ancestral relationships among genes; however, their analysis can be computationally prohibitive. We show here that these biological networks are compressible. They can be shrunk dramatically by eliminating redundant evolutionary relationships and this process is efficient because in these networks the number of compressible elements rises linearly rather than exponentially as in other complex networks. Compression enables global network analysis to computationally harness hundreds of interconnected genomes and to produce functional predictions. As a demonstration, we show that the essential, but functionally uncharacterized Plasmodium falciparum antigen EXP1 is a membrane glutathione S-transferase. EXP1 efficiently degrades cytotoxic hematin, is potently inhibited by artesunate, and is associated with artesunate metabolism and susceptibility in drug-pressured malaria parasites. These data implicate EXP1 in the mode of action of a frontline antimalarial drug.

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Section: Resultssupporting

confidence: 69%

Supergenomic Network Compression and the Discovery of EXP1 as a Glutathione Transferase Inhibited by Artesunate

Lisewski

Quiros

et al. 2014

Cell

113

128

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“…The formation of such a complex has been well studied by several investigators and now similar data are available for isozymes 3-3 and 4-4 from rat liver (Graminski et al 1989), rat liver microsomal GST (Weinander, Anderson, and Morgenstern 1994;Andersson et al 1995), class mu GST (Xinhua, Armstrong, and Gilliland 1993), human GSTA1-1 (Widersten, Bjornestedt, and Mannervik 1996;Hansson, Widersten, and Mannernik1997), as well as for pGSTP1-1 from porcine lung (Bico et al 1994). The results reported here with several model substrates indicate that the complex formation with the nitrobenzenes tested may adversely in uence the metabolism of endobioticsand xenobioticsvia the GST pathway in mammalian tissues.…”

Section: Discussionmentioning

confidence: 76%

In Vitro Inhibition of Mammalian Glutathione Transferases by Selected Nitrobenzenes

2000

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Five structurally related nitrobenzenes (1,2-dinitrobenzene, 1,3-dinitrobenzene, 1,4-dinitrobenzene, 1,3,5-trinitrobenzene, and picric acid) and Meisenheimer complex [1-(S-glutathionyl)-2,4,6-trinitrocyclohexadienate] were evaluated as possible inhibitors of af nity puri ed mammalian glutathione transferases (GSTs) isolated from human liver or human term placenta and rat liver. The results suggest that the degree of GST inhibition depends upon both the chemical in question and the enzyme source. Among the nitrobenzenes tested, 1,3,5-trinitrobenzene (TNB) was found to be the most potent inhibitor of GST activity toward 1-chloro-2,4-dinitrobenzene (CDNB) from all the sources, whereas 1,3-dinitrobenzene (1,3-DNB) was the least effective. TNB-caused inhibition of GST activity toward CDNB appeared to be isozyme speci c in that compared to the enzyme from human term placenta (GSTP1-1), the degree of inhibition of the mixture of GST isozymes present in the livers of adult rats and humans was low. The enzyme assays conducted with 3,4-dichloro-1-nitrobenzene (DCNB), ethacrynic acid (EA), and 4-nitropyridine N-oxide also suggested the isozymespeci c inhibition of rat liver GST activity by TNB. The nature of TNB-caused inhibition of GSTP1-1 was competitive with respect to CDNB and yielded a K i value of 12.5 µ M. With EA, a speci c substrate for GSTP1-1, an IC 50 value of » 16 µ M was estimated for the GSTP1-1 inhibition by TNB. The Meisenheimer complex, the product of nonenzymatic GSH conjugation with TNB by different GSTs, was found to be the most potent inhibitor of mammalian GSTs, and IC 50 values ranged between 1 and 4 µ M when the enzyme activity was assayed using CDNB. The nature of GSTP1-1 inhibition was noncompetitive with respect to CDNB, with a K i value of 1 µ M for Meisenheimer complex. Although a precise mechanism was not identi ed, it is postulated that GSH depletion and/or GST inhibition may contribute, at least partly, to the target organ toxicity caused by exposures of animals to different nitrobenzenes reported in the literature.

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“…In order to evaluate the global mechanism of MGST1 we have derived a steady-state rate equation for a random sequential kinetic mechanism for which there is experimental support 32 (Scheme 2, Equation 2).

v = \frac{k_{2} k_{3} false[normalC false] false[GSH false] {false[normalE false]}_{t}}{k_{- 2} normalX + k_{3} normalY + k_{2} normalZ}

normalX = K_{C} false[GSH false] + K_{C} K_{G} + false[normalC false] false[GSH false] + K_{G} false[normalC false] normalY = false[normalC false] false[GSH false] + K_{G} false[normalC false] normalZ = false[normalC false] false[GSH false] + K_{C} false[GSH false]

…”

Section: Resultsmentioning

confidence: 99%

“…To fill this gap we have performed a steady-state analysis of MGST1 at 5°C in order to determine whether the microscopic rate constants obtained for MGST1 accurately predict the steady-state kinetic parameters. The K M , k cat and k cat /K M values were also calculated from the steady-state rate equation for a random sequential mechanism 32 . The comparison between calculated and experimentally obtained kinetic parameters yields a reasonable fit for more reactive substrates and activated enzyme.…”

Section: Introductionmentioning

confidence: 99%

Global Kinetic Mechanism of Microsomal Glutathione Transferase 1 and Insights into Dynamic Enzyme Activation

et al. 2017

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Microsomal glutathione transferase 1 (MGST1) has a unique ability to be activated, up to 30-fold, by modification with sulfhydryl reagents. MGST1 exhibits one-third-of-the-sites-reactivity towards glutathione and hence heterogeneous binding to different active sites in the homo-trimer. Limited turnover stopped flow kinetic measurements of the activated enzyme allowed us to more accurately determine KD for the “third” low affinity GSH-binding site (1.4 ± 0.3 mM). The rate of thiolate formation, k2 (0.77 ± 0.06 s−1), relevant to turnover, could also be determined. By deriving the steady-state rate equation for a random sequential mechanism for MGST1 we can predict KM, kcat and kcat/KM values from these and previously determined pre steady-state rate constants (all determined at 5°C). To assess whether the pre steady-state behavior can account for the steady-state kinetic behavior we have determined experimental values for kinetic parameters at 5°C. For reactive substrates and activated enzyme, data for the microscopic steps account for the global mechanism of MGST1. For unactivated enzyme and more reactive electrophilic substrates, pre steady-state and steady-state data can be reconciled only if a more active subpopulation of MGST1 is assumed. We suggest that unactivated MGST1 can be partially activated in its unmodified form. The existence of an activated subpopulation (approximately 10%) could be demonstrated in limited turnover experiments. We therefore suggest that MSGT1 displays a pre-existing dynamic equilibrium between high and low activity forms.

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Kinetic studies on rat liver microsomal glutathione transferase: consequences of activation

Cited by 23 publications

References 26 publications

Supergenomic Network Compression and the Discovery of EXP1 as a Glutathione Transferase Inhibited by Artesunate

Supergenomic Network Compression and the Discovery of EXP1 as a Glutathione Transferase Inhibited by Artesunate

In Vitro Inhibition of Mammalian Glutathione Transferases by Selected Nitrobenzenes

Global Kinetic Mechanism of Microsomal Glutathione Transferase 1 and Insights into Dynamic Enzyme Activation

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