Subunit selective bleaching of C-phycocyanin from Synechococcus PCC 6301 strain AN 112 by Cu 2+ and sodium perchlorate

Abstract

The bleaching of the monomer and the separated α and β subunits of C-phycocyanin from Synechococcus PCC 6301 Strain AN 112 by Cu 2+ has been investigated. Upon the addition of Cu 2+ , the visible absorption band of the αβ monomer of phycocyanin was bleached and the maximum of the band shifted from 614 nm to 620 nm. The change in absorption induced by Cu 2+ was accompanied by a decreased fluorescence emission intensity and a shift of the fluorescence maximum from 649 nm to 643 nm. The difference spectra showed negative peaks at 604 nm in absorption and at 653 nm in fluorescence when the concentration of Cu 2+ was low. Spectroscopic titration curves showed biphasic behavior with respect to the concentration of Cu 2+ . Comparison of these results with the properties of the separated α and β subunits indicates that the β-subunit has higher affinity for Cu 2+ and is bleached preferentially over the α-subunit. This enables one to deduce the spectra of individual subunits from the monomer without separating them. This work also indicates that the β-subunit of phycocyanin is more labile than the α-subunit to denaturation with NaClO 4 . Partial unfolding of the protein in the presence of NaClO 4 appears to decrease the protein-chromophore interaction, allowing binding of Cu 2+ to the nitrogen atoms of the tetrapyrrole chromophores. This conclusion is supported by the results of a spectroscopic titration of biliverdin with Cu 2+ .