Phycoerythrin 545: Monomers, energy migration, bilin topography, and monomer/dimer equilibrium

Phycoerythrin 545 was isolated having an α₂β₂ (dimer) protein structure at pH 6.0 and 2 g/L protein concentration with eight bilin chromophores. Monomers (αβ) were produced by lowering the protein concentration to 0.15 g/L and the pH to 4.5. Dimer dissociation was monitored by dynamic light scattering and gel-filtration column chromatography. Monomers were stable and had bilin optical spectra different from the α₂β₂ dimers, although they have very similar protein secondary structures. The optical spectra of phycoerythrin 545 showed four types of behavior with temperature: 10-20 °C, dimers; 40-50 °C, dimers/monomers; 60 °C, nearly fully disordered; 70 °C, disordered α and β polypeptides. At 40 °C, the protein dissociated partially to monomer, which could be totally reversed to dimers at 20-25 °C. The visible circular dichroism difference spectrum for the protein dimers minus monomers exhibited positive and negative bands-such spectra may indicate exciton splitting between closely- spaced bilins. Circular dichroism also revealed a spectrum suggesting exciton coupling for the second excited state of the bilins. Ultrafast fluorescence using a two-photon method showed the fastest time for protein dimers to be 2.4 ps and monomers had a 39-ps lifetime. Phycocyanin 645 was found to have a 550-fs lifetime.