Conversion of γ-glutamylcysteinylethyl ester to glutathione in rat hepatocytes

Abstract

The conversion of gamma-glutamylcysteinylethyl ester (gamma-GCE) to glutathione in a reduced form (GSH) was examined using isolated rat hepatocytes pretreated with diethylmaleate, a GSH-depletor. Incubation of hepatocytes with 0.1 and 5.0 mM gamma-GCE (gamma-GCE-hepatocytes) over a 30-min period resulted in time-dependent increases in intracellular GSH and nonprotein-SH (NP-SH) concentrations. Hepatocytes incubated with 5.0 mM but not 0.1 mM GSH over a period of 30 min showed a time-dependent increase in intracellular GSH concentration. In the gamma-GCE-hepatocytes pretreated with bis-(p-nitrophenyl)phosphate (BNPP), a non-specific esterase inhibitor, an enhancement of intracellular GSH concentration was markedly reduced. gamma-GCE concentration in the gamma-GCE-hepatocytes with BNPP pretreatment was significantly higher than that in the cells without BNPP pretreatment, although there was no difference in the total amount of intracellular NP-SH, i.e., gamma-GCE, GSH, gamma-glutamylcysteine, cysteine ethyl ester, and cysteine between both gamma-GCE-hepatocytes. The present results indicate that gamma-GCE is transported into liver cells more easily than GSH itself, resulting in its conversion to GSH via esterase and glutathione synthetase within the cells.

Glutamine drives glutathione synthesis and contributes to radiation sensitivity of A549 and H460 lung cancer cell lines

“…The importance of glutamine-derived GSH for cell viability and against cytotoxicity was investigated using media supplemented with GSHE, the bioavailable form of GSH [2,6,27,29]. Initial treatment with 10 μM BPTES reduced viability and induced cytotoxicity (Figure 7a and 7b) in both tumor cell lines.…”

Mechanism of Protection by S-(1,2-Dicarboxyethyl) glutathione Triester against Acetaminophen-Induced Hepatotoxicity in Rat Hepatocytes.

“…Nishida et al elucidated the mechanism of conversion of gglutamylcysteinylethyl ester to GSH using isolated rat hepatocytes. 21) They reported that g-glutamylcysteinylethyl ester was transported into liver cells more easily than GSH itself, resulting in its conversion to GSH via esterase and glutathione synthetase within the cells, and also reported that the elevation of the intracellular GSH concentration induced on treatment with g-glutamylcysteinylethyl ester was markedly reduced by treatment with bis-(p-nitrophenyl)phosphate. From this evidence it was assumed that the DCE-GS triester transported into hepatocytes was hydrolyzed to DCE-GS via the di-and monoester by esterase.…”

Mechanistic role of each metal ion in Streptomyces dinuclear aminopeptidase: Peptide hydrolysis and 7×1010-fold rate enhancement of phosphodiester hydrolysis

“…Moreover, BNPP and a few phosphoesters and fluorophosphates are also known to be TS à -inhibitors against serine proteases and esterases such as trypsin, chymotrypsin, and acetylcholine esterase through covalent linkage with the nucleophilic serine in the active site [7,[11][12][13] to afford Ser-O-P(O) 2 --OR with concomitant cleavage of a phosphoester or the P-F bond that resembles the ''initial burst" kinetics of serine proteases. However, the phosphoester Ser-O-P(O) 2 --OR is indefinitely stable, thus has only one turn-over for the phosphoester bond cleavage by these enzymes.…”

“…Moreover, the phosphoester bis(p-nitrophenyl)phosphate (BNPP) and the phosphonate ester p-nitrophenyl phenylphosphonate can be effectively hydrolyzed by the dinuclear aminopeptidase (AP) from Streptomyces griseus (SgAP) with activities comparable to some native phosphoesterases [8][9][10]. Since phospho-and phosphono-esters are TS à -like molecules and can inhibit peptide hydrolysis [11][12][13], their hydrolysis by SgAP is novel and must take place according to a unique catalytic pathway of the enzyme.…”