MONOMERIC C-PHYCOCYANIN AT ROOM-TEMPERATURE AND 77-K - RESOLUTION OF THE ABSORPTION AND FLUORESCENCE-SPECTRA OF THE INDIVIDUAL CHROMOPHORES AND THE ENERGY-TRANSFER RATE CONSTANTS

At both room temperature (RT) and 77 K, the absorption and fluorescence spectra of the three individual chromophore types (alpha84, beta84, and beta155) found in monomeric C-phycocyanin (alpha(PC)beta(PC)), isolated from the cyanobacterium Synechococcus sp. PCC 7002, were resolved along with the rates of energy transfer between the chromophores. The cpcB/C155S mutant, whose PC is missing the beta155 chromophore, was useful in effecting this resolution. At RT, the single broad peak in the visible region of the absorption spectrum of (alpha(PC)beta(PC)) was resolved into its three-component spectra by comparing the steady-state absorption spectra of the isolated wild-type alpha subunit of PC (alpha(PC)) (containing only the alpha84 chromophore) with those of the monomeric PCs isolated from the mutant strain (alpha(PC)beta*) and the wild-type strain (alpha(PC)beta*). At 77 K, the visible region of the absorption spectrum of (alpha(PC)beta(PC)) splits into two peaks. This partial resolution at 77 K of the chromophore spectra of (alpha(PC)beta(PC)) when compared with the 77 K absorption spectra of alpha(PC), beta(PC), and (alpha(PC)beta*) provided a confirmation of our RT assignments of the chromophore absorption spectra. The individual fluorescence spectra of the chromophores and the rate constants for energy transfer between them in (alpha(PC)beta(PC)) were resolved by modeling the time-resolved fluorescence spectra of beta(PC)(containing the beta155 and beta84 chromophores) and (alpha(PC)beta*). As with absorption, lowering the temperature to 77 K helped resolve the fluorescence spectra of the individual chromophores because the spectra were narrowed.