Studies on plantacyanin. IV. Reconstitution with Cu-thionein, oxidation by cytochrome oxidase and autooxidation in the presence of cardiolipin.

Cu-thionein isolated from cucumber roots was used for reconstitution of plantacyanin from cucumber. The rate of the copper transfer from Cu-thionein to apoplantacyanin was found to depend on pH, ionic strength and concentrations of the proteins. The rate of reconstitution with Cu-thionein was 10-times higher than with copper ions. No intermediate was observed during reconstitution with Cu-thionein. The incubation of oxidized holoplantacyanin with Cu-thionein or apothionein brings about the reduction of plantacyanin copper. This process, however, was found to be slow as compared to the rate of copper transfer from Cu-thionein to apoplantacyanin. Cytochrome oxidase from heart mitochondria was detected to possess some plantacyanin oxidase activity with the turnover number 5 min-1. The activity of the enzyme towards plantacyanin as well as with cytochrome c as a substrate was established to be lipid and ionic strength-dependent, and it was inhibited by CN- and N3-. Lineweaver-Burk plots show that the inhibitory effect of ionic strength on plantacyanin oxidase activity is connected with changes of Michaelis constant rather than of the maximal rate. Plantacyanin which is known to be very resistant towards many cationic, anionic and nonionic detergents, becomes, as well as cytochrome c, autooxidable in the presence of cardiolipin.