Kinetic analysis of nonphotochemical quenching of chlorophyll fluorescence. 2. Isolated light-harvesting complexes.

The chlorophyll fluorescence yield of purified photosystem II light-harvesting complexes can be lowered by manipulation of experimental conditions. In several important respects, this quenching resembles the nonphotochemical quenching observed in isolated chloroplasts and leaves, therefore providing a model system for investigating the underlying mechanism. A methodology based on the principles of enzyme kinetic analysis has already been applied to isolated chloroplasts, and this same experimental approach was used here with purified LHCIIb, CP26, and CP29. It was found that the kinetics of the decrease in fluorescence yield robustly fitted a second-order kinetic model with respect to time after induction of quenching. The second-order rate constant was dependent upon the complex that was analyzed, the detergent concentration, the solution pH, and the presence of exogenous xanthophyll cycle carotenoids. In contrast, the formation of an absorbance change at 683 nm that accompanies quenching displayed first-order kinetics. The reversal of quenching also displayed second-order kinetics. These data show that quenching results from a binary reaction, possibly arising between two chlorophyll molecules. On the basis of these data, a model for the regulation of nonphotochemical quenching based upon the allosteric control of the conformation of light-harvesting complexes by protonation and xanthophyll binding is presented.