Proteasome inhibition in glyoxal-treated fibroblasts and resistance of glycated glucose-6-phosphate dehydrogenase to 20 S proteasome degradation in vitro.
Glycation and glycoxidation protein products are formed upon binding of sugars to NH(2) groups of lysine and arginine residues and have been shown to accumulate during aging and in pathologies such as Alzheimer's disease and diabetes. Because the proteasome is the major intracellular proteolytic system involved in the removal of altered proteins, the effect of intracellular glycation on proteasome function has been analyzed in human dermal fibroblasts subjected to treatment with glyoxal that promotes the formation of N epsilon-carboxymethyl-lysine adducts on proteins. The three proteasome peptidase activities were decreased in glyoxal-treated cells as compared with control cells, and glyoxal was also found to inhibit these peptidase activities in vitro. In addition, the activity of glucose-6-phosphate dehydrogenase, a crucial enzyme for the regulation of the intracellular redox status, was dramatically reduced in glyoxal-treated cells. Further analysis was performed to determine whether glycated proteins are substrates for proteasome degradation. In contrast to the oxidized glucose-6-phosphate dehydrogenase, both N epsilon-carboxymethyl-lysine- and fluorescent-glycated enzymes were resistant to degradation by the 20 S proteasome in vitro, and this resistance was correlated with an increased conformational stability of the glycated proteins. These results provide one explanation for why glycated proteins build up both as a function of disease and aging. Finally, N epsilon-carboxymethyl-lysine-modified proteins were found to be ubiquitinated in glyoxal-treated cells suggesting a potential mechanism by which these modified proteins may be marked for degradation.
Bulteau
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AL
Laboratoire de Biologie et Biochimie Cellulaire du Vieillissement, Université Denis Diderot, Paris 7, 2 Place Jussieu, CC 7128, 75251 Paris Cedex 05, France.
Verbeke
P
P
Petropoulos
I
I
Chaffotte
A F
AF
Friguet
B
B
D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000187:drug effects*; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D005347:Fibroblasts / Q000187:drug effects; D005456:Fluorescent Dyes / Q000378:metabolism; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism*; D017127:Glycation End Products, Advanced / Q000378:metabolism*; D006037:Glyoxal / Q000494:pharmacology*; D006801:Humans; D006868:Hydrolysis; D009097:Multienzyme Complexes / Q000187:drug effects*; D046988:Proteasome Endopeptidase Complex; D013050:Spectrometry, Fluorescence
D000817:Anilino Naphthalenesulfonates; D015853:Cysteine Proteinase Inhibitors; D005456:Fluorescent Dyes; D017127:Glycation End Products, Advanced; D009097:Multienzyme Complexes; D006037:Glyoxal; C027132:1-anilino-8-naphthalenesulfonate; D005954:Glucosephosphate Dehydrogenase; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex