GSH/GSSG ratios

Glutathione reductase (GR) catalyzes the reduction of oxidized glutathione (GSSG) to reduced glutathione (GSH) using NADPH provided from the hexose monophosphate pathway. GR, a ubiquitous flavoenzyme, maintains a high value of two for the GSH/GSSG ratio in the red blood cells. l,3-Bis(2-chloroethyl)-nitrosourea (BCNU) selectively inhibits cellular GR. GR is composed of two identical subunits, each of molecular mass 50 kDa (S8). The three-dimensional structure and mechanism of catalysis have been established for human GR (K17). 

III. Glutathione reductase (EC 1.6.4.2) It is a flavoprotein that catalyzes the NADPH-dependent reduction of oxidized glutathione (GSSG) to glutathione (GSH). This enzyme is essential for the GSH redox cycle which maintains adequate levels of reduced cellular GSH. A high GSH/GSSG ratio is essential for protection against oxidative stress. 

The reduced form of Na+, K+-ATPase inhibitor-I (10) was obtained by treatment of the protected peptide synthesized by the soln procedure with HF, followed by reaction with Hg(OAc)2. After purification of the crude product on Sephadex G-25, the reduced peptide (110 mg) was dissolved in 0.1 M NH4OAc buffer (1L, pH 7.8) at a peptide concentration of 0.018 mM and then stirred at rt. After 24 h, the major peak in the HPLC, which coeluted with the natural product, corresponded to 55% of the product distribution. The mixture was acidified to pH 3 with AcOH and 10 was purified by RP-HPLC. When the oxidation was carried out in the presence redox reagents at a peptide/GSH/GSSG ratio of 1 100 10, after 24 h the major oxidation product increased to 69%. The mixture was acidified with AcOH and the product (10) isolated by preparative HPLC yield 20%. The product was characterized by MALDI-TOF-MS and amino acid analysis a combination of enzymatic peptide mapping and synthetic approaches were applied to assign the cystine connectivities. 

As exemplarily shown in the case of charybdotoxin, a 37-residue peptide with three intramolecular disulfide bonds,[70] operating in redox buffer was crucial for efficient formation of the correct disulfide bonds.[71] When the reduced peptide was oxidized in 0.1 M NHtOAc buffer (pH 8.0) at 0.11 mM concentration in the presence of redox reagents (peptide/GSH/GSSG ratio of 1 60 6), the main product was the native peptide contaminated... 

Very dynamic glutathione redox cycle activity to maintain GSH/GSSG ratio of about 50-100... 

Active glutathione redox cycle that maintains a GSH/GSSG ratio of about 25-50... 

Glutathione reductase catalyzes the virtually irreversible reduction of GSSG by NADPH. Its metabolic function is therefore synonymous with that of the product GSH. Glutathione is the most abundant thiol-disulfide pair in the cell by more than an order of magnitude under most conditions the GSH GSSG ratio is about 20 1 (195). Since the ratio of... 

Figure 2. The incubation in the presence of GSSG induces the S-glutathionylation of c-Jun and an inhibition in its DNA binding activity. A) The DNA binding of c-Jun was subjected to EMSA experiments in the presence of different GSH/GSSG ratios or DTT. B) In the figure is represented the amount of c-Jun protein which is modified to mixed disulfide with glutathione (P-S-S-G) or to other modifications.

Oxidative stress GSH-GSSG ratio, generation of free radical species, lipid peroxidation products and DNA strand breaks... 

Fig. 18. Effect of glucose on insulin release, glucose oxidation, pentose phosphate shunt, NADH/NAD +, NADPH/NADP"1", GSH/GSSG ratios and Ca + uptake of rat pancreatic islets. In a concentration range up to 16.7 mM, glucose-mediated stimulation of insulin secretion is closely parallelled by increases in glucose oxidation, pentose phosphate shunt activity, the NADH/NAD+, NADPH/NADP+, GSH/GSSG ratios and Ca2+ uptake (Ammon and Wahl, 1994).

If we accept that the redox state of nicotinamide nucleotides is a modulating system, we need to know how this works. An attractive idea involves the fact that the NADPH/NADP+ ratio is in equilibrium with the GSH/GSSG ratio which itself is of importance for the redox state and therefore the activity of functional proteins (Barron, 1951). 

Fig. 19. Effect of diazenedicarboxylic acid bis-(/V -methylpiperazide) (DIP) on insulin release, ATP concentration, GSH/GSSG ratio and Ca"+ uptake of rat pancreatic islets. DIP, an oxidant of GSH at the concentration used here, did not affect islet ATP concentration but inhibited glucose-induced increases in GSH/GSSG ratio, 45Ca2+ uptake and insulin release (Ammon and Wahl, 1994).

Previous evidence suggests that it is the change in the GSH/GSSG ratio rather than that in GSH levels as such that modulates insulin secretion buthionine-sulphoximine (BSO), an inhibitor of glutathione synthesis (Griffith and Meister, 1979), dramatically decreased islet GSH in rats when administered in vivo, but did not affect its GSH/GSSG ratio. Under these conditions, glucose-induced release of insulin was not affected (Klumpp and Ammon, 1988). 

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