Global target analysis of picosecond chlorophyll fluorescence kinetics from pea chloroplasts: A new approach to the characterization of the primary processes in photosystem II alpha- and beta-units.

In this study, we have used the method of target analysis to analyze the ps fluorescence kinetics of pea chloroplasts with open (F(0)) and closed (F(max)) photosystem II (PS II) centers. Extending the exciton/radical pair equilibrium model (Schatz, G. H., H. Brock, and A. R. Holzwarth. 1988. Biophys. J. 54:397-405) to allow for PS II heterogeneity, we show that two types of PS II (labeled alpha and beta) must be accounted for, each pool being characterized by its own set of molecular rate constants within the model. Simultaneous global target analysis of the data at F(0) and F(max) results in a detailed description of the molecular kinetics and energetics of the primary processes in both types of PS II units. This characterization revealed that the PS IIalpha pool accounts for twice as many Chl molecules as PS IIbeta, which suggests a PSIIalpha/PSIIbeta reaction center stoichiometry of close to unity. By extrapolation it is shown that the primary charge separation in hypothetical "isolated" beta reaction centers is slower than in isolated alpha reaction centers: in open centers by a factor of 4 (1/k(1) (int) = 11 vs 2.9 ps), in closed centers by a factor of 2 (1/k(1) (int) = 34 vs 19 ps). Despite this slower charge separation process in PS IIbeta, the quantum efficiency of the charge separation process is hardly affected: a charge stabilization yield at F(0), (i.e., P(+)IQ(A) (-)) of 86% (as compared to 90% in PS IIalpha). Reduction of Q(A) (closing PS II) has distinctly different effects on the primary kinetics of PS IIbeta, as compared to PS IIalpha. In PS IIalpha the charge separation rate drops by a factor of 6, whereas the charge recombination process is hardly affected. In PS IIbeta the charge separation is slowed down by a factor of 3, whereas the charge recombination rate increases by a factor of 5. In terms of changes in standard free energy, the reduction to Q(A) (-) lifts the free energy of the radical pair P(+)I(-), relative to the excited state (Chl(n)/P)(*), by 47 meV in PS IIalpha and by 67 meV in PS IIbeta. The concomitant increase in fluorescence quantum yield is the same for both types of PS II. These results show that PS IIalpha and PS IIbeta exhibit a different molecular functioning with respect to the primary processes, which might have its origin in a different molecular structure of the reaction centers and/or a different local environment of these centers. Location in different parts of the thylakoid membrane might be involved. We also applied different error analysis procedures to determine the error ranges of the values found for the molecular rate constants. It is shown that the commonly used standard error has very little meaning, as it assumes independence of the fit parameters. Instead, an exhaustive search procedure, accounting for all possible correlations between the fit parameters, gives a more realistic view on the accuracy of the fit parameters.