COMPARISON OF CALCULATED AND EXPERIMENTALLY RESOLVED RATE CONSTANTS FOR EXCITATION-ENERGY TRANSFER IN C-PHYCOCYANIN .2. TRIMERS

The light-harvesting protein C-phycocyanin (PC) in the trimeric aggregation state, isolated from the cyanobacterium Synechococcus sp. PCC 7002, is studied by absorption spectroscopy and by time-resolved anisotropic fluorescence spectroscopy with 1 ps time resolution. PC trimers isolated from the wild-type strain and a mutant strain, cpcB/C155S, in which the P155 chromophore is missing, are compared. The absorption spectra of the trimeric PCs, when compared with previously published spectra of the monomeric PCs [Debreczeny et al. J. Phys. Chem. 1993, 97, 9852-9862], lead us to conclude that the absorption spectrum of the beta(155) chromophore is similar when PC is in the monomeric and trimeric states. This means that the red shift of the absorption spectrum that occurs when PC aggregates from monomers to trimers is due to changes in the spectra of the alpha(84) and/or beta(84) chromophores. First-order exciton coupling between chromophores cannot alone be the cause of the red shift. Time-reserved fluorescence anisotropy measurements lead to assignments of the dominant rate constants for energy transfer between chromophores in PC trimers and provide information about the relative chromophore orientations. The angle between the transition dipoles of the alpha(84)(1) and beta(84)(2) chromophores on adjacent monomers in the trimer is estimated from the fluorescence anisotropy decay to be 52 degrees (the chromophore numbering scheme follows the convention established by Schirmer et al. J. Mol. Biol. 1987, 196, 677-695). The angles between the C-3 axis of symmetry in the trimer and the alpha(84) and beta(84) chromophore transition dipoles are restricted to four possible values. The observed anisotropic fluorescence decay times of 1.0, 50, and 40 ps are respectively assigned to energy transfer within the alpha(84)(1)-beta(84)(2) pairs on adjacent monomers, within the beta(155)(1)-beta(84)(1) pairs on the same monomer, and between the alpha(84)(1) beta(84)(2), alpha(84)(2) beta(84)(3), and alpha(84)(3) beta(84)(1) pairs around the trimer ring. These results are in excellent agreement with Forster calculations in the weak coupling limit which predict decay times of 1.4, 49, and 46 ps for these same three energy-transfer processes. In combination with previous results [Debreczeny et al., companion paper], these results indicate that the dominant energy-transfer processes in monomeric and trimeric PC are well described by Forster's theory. This is the first detailed confirmation of Forster theory in a pigment protein.