SummaryThe involvement of sulfhydryl groups in the Cu binding and reconstitution of apoceruloplasmin was investigated. Recombination studies in which different Cu reagents were added to the apoprotein demonstrated a preference of the apoprotein for Cu(I) generated in solution by the action of a suitable reductant. The -SH titer for apoproteins prepared by different methods was correlated with the percentage reconstitution obtained in the Cu-protein recombination studies. It was found that reconstitution of the apoprotein was proportional to the average number of -SH groups/molecule in the protein preparation. The results of this study suggest that apoceruloplasmin contains 4 -SH groups/molecule and that these -SH groups are necessary for reconstitution. Under our conditions, native ceruloplasmin was found to have less than 0.5 -SH groups/molecule.