Oxidative-stress induced protein glutathione mixed-disulfide formation in the ocular lens.
Biochim Biophys Acta
; 1313(1): 20-8, 1996 Aug 21.
Article
in En
| MEDLINE
| ID: mdl-8781545
The biochemistry of protein-glutathione mixed disulfide formation in the ocular lens was examined by 13C-NMR spectroscopic measurements of glutathione oxidative metabolism in intact rabbit lenses maintained in organ culture. Lenticular amino acid uptake and glutathione biosynthetic mechanisms were employed to facilitate the incorporation of L-[3-13C]cysteine from the incubation medium into the cysteinyl residue of glutathione. Subsequent exposure to increasing levels of oxidative stress induced by tert-butylhydroperoxide resulted in decreased levels of ([3-13C]cysteinyl)-glutathione and a loss of 13C NMR resonance intensity, a reflection of protein-glutathione mixed disulfide formation. The rate of ([3-13C]cysteinyl)-glutathione loss depended on the concentration of tert-butylhydroperoxide; 13C-labeled oxidized glutathione was observed only at the highest concentration (2 mM) of oxidant tested. Removal of the oxidative stress led to a partial recovery of ([3-13C]cysteinyl)-glutathione levels and 13C resonance intensity. Recovery was significantly enhanced by the addition of 2-mercaptoethanol. The mechanism of protein-glutathione adduct formation was further characterized by the in vitro monitoring of the reaction of oxidized glutathione with bovine lens gamma-II crystallin protein using proton NMR spectroscopy. These experiments provided insight into the role of the cellular glutathione redox-couple, [GSH]/[GSSG], in maintaining reduced protein thiol groups, and suggested that protein-glutathione adduct formation may function as a mechanism for modulating the glutathione redox buffer under conditions of oxidative stress in ocular tissue. In addition, the results demonstrate the feasibility of direct chemical reduction of protein-glutathione disulfide bonds in vivo which may reflect a mechanism for the inhibition of disulfide-linked light scattering protein aggregate formation.
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Collection:
01-internacional
Database:
MEDLINE
Main subject:
Oxidative Stress
/
Glutathione
/
Lens, Crystalline
Limits:
Animals
Language:
En
Journal:
Biochim Biophys Acta
Year:
1996
Document type:
Article
Affiliation country:
United States
Country of publication:
Netherlands