Cu2+-catalyzed oxidative degradation of thyroglobulin.
Thyroglobulin (Tg) was subjected to metal-catalyzed oxidation, and the oxidative degradation was analyzed by SDS-polyacrylamide gel electrophoresis under reducing conditions. In contrast to no effect of hydrogen peroxide (H2O2) alone on the Tg degradation, the inclusion of Cu2+ (30 microM), in combination with 2 mM H2O2, caused a remarkable degradation of Tg, time- and concentration-dependent. The action of Cu2+ was not mimicked by Fe2+, suggesting that Tg may interact selectively with Cu2+. A similar degradation of Tg was also observed with Cu2+/ascorbate system, and the concentration of Cu2+ (5-10 microM), in combination with ascorbate, required for the effective degradation was smaller than that of Cu2+ (10-30 microM) in combination with H2O2. In support of involvement of H2O2 in the Cu2+/ascorbate action, catalase expressed a complete protection. However, hydroxyl radical scavengers such as dimethylsulfoxide or mannitol failed to prevent the oxidation of Tg whereas phenolic compounds, which can interact with Cu2+, diminished the oxidative degradation, presumably consistent with the mechanism for Cu2+-catalyzed oxidation of protein. Moreover, the amount of carbonyl groups in Tg was increased as the concentration (3-100 microM) of Cu2+ was enhanced, while the formation of acid-soluble peptides was not remarkable in the presence of Cu2+ up to 200 microM. In further studies, Tg pretreated with heat or trichloroacetic acid seemed to be somewhat resistant to Cu2+-catalyzed oxidation, implying a possible involvement of protein conformation in the susceptibility to the oxidation. Based on these observations, it is proposed that Tg could be degraded non-enzymatically by Cu2+-catalyzed oxidation.
Lee
H J
HJ
College of Pharmacy, Chungnam National University, Taejon, Korea.
Sok
D E
DE
D000818:Animals; D001205:Ascorbic Acid / Q000737:chemistry; D002374:Catalase / Q000494:pharmacology; D003300:Copper / Q000008:administration & dosage / Q000737:chemistry*; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D016166:Free Radical Scavengers; D006358:Hot Temperature; D006861:Hydrogen Peroxide / Q000008:administration & dosage / Q000737:chemistry; D017665:Hydroxyl Radical; D007700:Kinetics; D008353:Mannitol / Q000494:pharmacology; D016877:Oxidants / Q000737:chemistry*; D010084:Oxidation-Reduction; D010636:Phenols / Q000494:pharmacology; D013552:Swine; D013954:Thyroglobulin / Q000737:chemistry*; D014238:Trichloroacetic Acid / Q000494:pharmacology
D016166:Free Radical Scavengers; D016877:Oxidants; D010636:Phenols; D017665:Hydroxyl Radical; D008353:Mannitol; D014238:Trichloroacetic Acid; D003300:Copper; D013954:Thyroglobulin; D006861:Hydrogen Peroxide; D002374:Catalase; D001205:Ascorbic Acid; D004121:Dimethyl Sulfoxide