Deglycosylated ceruloplasmin maintains its enzymatic, antioxidant, cardioprotective, and neuronoprotective properties.
Ceruloplasmin (CP), an important serum antioxidant, is a blue copper glycoprotein with ferroxidase and oxidase activities. Among other physiological actions, plasma CP was shown to protect isolated rat hearts and cultured P19 neurons exposed to oxidative stress conditions, raising the possibility of using this protein in the treatment of cardiac and neuronal diseases related to oxidative damage. However, since therapeutic applications of CP must be compatible with restrictions in the administration of blood derivatives to humans, there is a need to produce the protein by genetic engineering. To help in the choice of adequate expression systems, we undertook this study to determine if the carbohydrate moiety on the protein is essential for its functions. CP was completely deglycosylated using N-glycosidase F under nondenaturing conditions. Deglycosylated CP was found to retain most of the conformational, antioxidant, and enzymatic properties of the native protein in vitro. Moreover, both forms of the protein had similar cardioprotective and neuronoprotective effects against oxidative stress as evaluated with isolated rat hearts undergoing ischemia-reperfusion and with cultured P19 neurons exposed to xanthine-xanthine oxidase. The data thus indicate that the carbohydrate moiety of CP is not essential for its enzymatic and protective actions. Accordingly, even the use of expression systems that do not glycosylate mammalian proteins could provide a recombinant CP that retains its therapeutic potential.
Aouffen
M
M
Department of Chemistry and Biochemistry, Université du Quebec à Montreal, Succ. Centre-ville, Canada.
Paquin
J
J
De Grandpré
E
E
Nadeau
R
R
Mateescu
M A
MA
D000581:Amidohydrolases / Q000378:metabolism; D000975:Antioxidants / Q000378:metabolism*; D002316:Cardiotonic Agents / Q000378:metabolism*; D002460:Cell Line; D002570:Ceruloplasmin / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D018696:Neuroprotective Agents / Q000378:metabolism*; D018384:Oxidative Stress; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
D000975:Antioxidants; D002316:Cardiotonic Agents; D018696:Neuroprotective Agents; D002570:Ceruloplasmin; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase