Alteration of enzymes in ageing human fibroblasts in culture. V. Mechanisms of glutathione peroxidase modification.
Ageing of WI-38 fibroblasts in culture was used as a model in order to investigate the evolution and the alteration of the key antioxidant enzyme glutathione peroxidase. The activity of glutathione peroxidase is influenced by the presence of selenium in the culture medium and we have also shown that the specific activity of this enzyme does not decrease during ageing, but rather slightly increases. No alteration could be detected by immunotitration. Also the kinetic parameter Km for tert-butyl hydroperoxide has not changed. However, the heat resistance of the enzyme dramatically decreases with ageing. Dilutions of the enzyme preparations had the same influence on the thermosensitivity of the enzyme. This dilution effect is most probably linked to the dissociation of the enzyme subunits into dimers and monomers. Moreover, the kinetic of thermoinactivation curves are best explained by consecutive reactions of inactivation with an intermediary enzyme form. These observations strongly support the hypothesis that ageing is associated with an increased dissociation constant of the tetrameric glutathione peroxidase leading to an easier dissociation of the enzyme in old cells.
Pigeolet
E
E
Facultés Universitaires N.-D. de la Paix, Namur, Belgium.
Remacle
J
J
D002460:Cell Line; D002470:Cell Survival; D002478:Cells, Cultured; D003470:Culture Media; D004789:Enzyme Activation; D000939:Epitopes; D005347:Fibroblasts / Q000201:enzymology*; D005979:Glutathione Peroxidase / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D009994:Osmolar Concentration; D012645:Selenomethionine / Q000494:pharmacology; D013696:Temperature
D003470:Culture Media; D000939:Epitopes; D012645:Selenomethionine; D005979:Glutathione Peroxidase