Spectral analysis of pigment photobleaching in photosynthetic antenna complex LHCIIb

Light-induced photooxidation of chlorophyll (Chl) a, b and xanthophylls was investigated in LHCIIb, the antenna pigment–protein complex of photosystem II. Absorption difference spectra at normal and low temperatures show initially (at less than 25% Chl a decay) a selective bleaching of a red-shifted Chl b with absorption bands at 487 and 655 nm, Chl b (460/650 nm) and Chl a (433/670 nm), which changes to a less selective photooxidation pattern at deeper bleaching stages. Difference absorption spectra and HPLC analyses indicate different photooxidation rates of pigments in the order neoxanthin>Chl a>lutein≈Chl b. Despite significant pigment loss as monitored with absorption spectra, CD spectra indicate an essentially complete persistence of the protein secondary structure. Fluorescence excitation spectra suggest the conversion of a small fraction of Chl a into pheophytin a which acts as a fluorescence quencher, possibly through temporary charge separation process. The strong features in the electroabsorption (Stark effect) spectra due to chlorophyll b at 655 nm and a xanthophyll at 510 nm, and the spectral changes mentioned above are assigned to Chl molecules located at several binding sites in LHCIIb protein and are discussed in the context of spatial configuration and interactions of pigment molecules.