Gulonolactone oxidase activity-dependent intravesicular glutathione oxidation in rat liver microsomes

• Published in: FEBS letters 1998-07, Vol.430 (3), p.293-296
• Main Authors: Puskás, Ferenc, Braun, László, Csala, Miklós, Kardon, Tamás, Marcolongo, Paola, Benedetti, Angelo, Mandl, József, Bánhegyi, Gábor
• Format: Article
• Language: English
• Subjects: Alamethicin - pharmacology; Animals; Ascorbate; Ascorbic Acid - metabolism; DIDS, 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid; Endoplasmic reticulum; Enzyme Activation; Glutathione; Glutathione - metabolism; Glutathione Disulfide - metabolism; GSH, glutathione (reduced form); GSSG, glutathione (oxidised form); Gulonolactone oxidase; L-Gulonolactone Oxidase; Light; Male; Microsomes, Liver - enzymology; MOPS, 4-morpholinepropanesulfonic acid; Oxidation-Reduction; PEG, polyethylene glycol; Rats; Rats, Sprague-Dawley; Scattering, Radiation; Sugar Acids - metabolism; Sugar Alcohol Dehydrogenases - metabolism; Uncoupling Agents - pharmacology
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/S0014-5793(98)00678-4

The orientation of gulonolactone oxidase activity was investigated in rat liver microsomes. Ascorbate formation upon gulonolactone addition resulted in higher intravesicular than extravesicular ascorbate concentrations in native microsomal vesicles. The intraluminal ascorbate accumulation could be prevented or the accumulated ascorbate could be released by permeabilising the vesicles with the pore-forming alamethicin. The formation of the other product of the enzyme, hydrogen peroxide caused the preferential oxidation of intraluminal glutathione in glutathione-loaded microsomes. In conclusion, these results suggest that the orientation of the active site of gulonolactone oxidase is intraluminal and/or the enzyme releases its products towards the lumen of the endoplasmic reticulum.