Age-related changes in human ceruloplasmin. Evidence for oxidative modifications.
Human plasma or serum from donors of age comprised between 15 and 95 years was analyzed for paramagnetic and total copper content, as well as for immunoreactive ceruloplasmin content and oxidase activity. All parameters were essentially unaltered, except the paramagnetic copper content, which increased 2-fold upon aging. A dramatic change of the electron paramagnetic resonance spectrum due to ceruloplasmin occurred in individuals over 65 years old and was associated with both an increase of the type 1 copper signal intensity and the appearance of new resonances of a type 2 copper species. Ceruloplasmin was isolated from either young or old donors. Spectroscopic analyses of the isolated proteins confirmed the tendency of type 1 copper to stay reduced in the "young" and oxidized in the "old" protein. The type 2 copper signal observed in most young ceruloplasmin samples was different from the species invariably present in the old protein. The magnetic parameters of the latter species were more consistent with a partially reduced trinuclear copper site. In vitro limited proteolysis resulted in identical fragmentation patterns and kinetics in both proteins. However, changes of the net electric charge were detected in the fragments of the protein isolated from aged individuals, which exhibited a carbonyl content of 0.6 mol of carbonyl/mol of protein. The same pattern of modifications, including a higher carbonyl content (0.65 versus 0.2 mol of carbonyl/mol of protein), could be reproduced by exposure of the young protein to the metal-catalyzed oxidation system iron/ascorbate. These results suggest that during aging ceruloplasmin is subjected to oxidative modifications which are likely to be the source of conformational changes around the copper sites leading to an intramolecular electron rearrangement among the various copper sites.
Center of Molecular Biology of Consiglio Nazionale delle Ricerche, University of Rome La Sapienza, Italy.
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Musci
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MC
Bonaccorsi di Patti
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U
Fagiolo
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Calabrese
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000375:Aging / Q000097:blood*; D002570:Ceruloplasmin / Q000378:metabolism*; D003300:Copper / Q000097:blood; D004578:Electron Spin Resonance Spectroscopy; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D008875:Middle Aged; D010084:Oxidation-Reduction
D003300:Copper; D002570:Ceruloplasmin