Copper metalloproteins are among the primary oxidases, oxygenases and certain oxygen carriers in animal cells. The long range goal is the elucidation of the chemistry, function, metabolism and biosynthesis of Cu metalloproteins, with emphasis on ceruloplasmin (Cp), because it is central to the transport and metabolism of copper. The oxidation of cysteine represents a new class of thiol substrates for the oxidase action at pH 7 of Cp. It is proposed to make a thorough study of the structure-activity relationships for cysteine and other thiol substrates. High specificity is indicated by fact that 3,3-dimethylcysteine (penicillamine) is not oxidized. The products resulting from the Cp catalyzed oxidation of cysteine and derivatives will be studied using mass spectrometry or other methods as necessary. The mechanism of the catalytic oxidation of cysteine by Cp will also be investigated using a variety of physical and chemical methods. Our focus will be on the shift in pH optimum, the production of superoxide ion and the absence of inhibition by azide. Further study of the protective and anti-oxidant activity of Cp will involve the Cu(II) induced hemolysis of red blood cells as a model system and its prevention by Cp. Iron mobilization and transport, for which Cp is the molecular link between copper and iron metabolism, will be examined for the effect of serum factors, especially apotransferrin, albumin, biocarbonate and citrate. Cp-free serum will also be developed for this purpose. Catalytic and/or receptor mechanisms for the mobilization of iron by ceruloplasmin will be sought. The long term goals of this research are directed toward relating the structure of Cp to its multi-functions, determining how Cu ion reacts with apoceruloplasmin in vivo, and how Cu is released from this circulating copper protein to fulfill its transport function.
