Structural role of the copper ions in the dinuclear active site of Carcinus aestuarii hemocyanin.

In this work we show, by a combination of biochemical and biophysical approaches, that the copper ions bound in the binuclear active site of Carcinus aestuarii hemocyanin play a stabilizing role on the tertiary structure of the protein. Upon removal of copper, the monomeric hemocyanin, but not the hexameric oligomer, undergoes changes at the level of tertiary structure while the secondary structure is almost unaffected. By Small-Angle X-Ray Scattering, supported by gel chromatography measurements, it can be concluded that the apo-monomer, but not the holo form or the hexameric form, undergoes a slow time-dependent oligomerization process.