Two-Photon-Sensitized Fluorescence and Excitation Spectra of Photosystem I ofThermosynechococcuselongatus

Abstract

In this work, we present the two-photon excitation spectrum and two-photon-sensitized fluorescence spectra of photosystem I (PS I) of Thermosynechococcus elongatus. The two-photon excitation spectrum of PS I agrees well with known spectra of the carotenoid (Car) S1 state of β-carotene in solution. Only a small intensity of the PS I two-photon spectrum around its 0−0 transition indicates hot-state Car S1 → chlorophyll (Chl) energy transfer. The two-photon-sensitized fluorescence spectrum of PS I shows no major difference of the intensities of bulk- and red-chlorophyll fluorescence in comparison to the fluorescence spectrum observed after nonselective one-photon excitation of the PS I chlorophylls. Fluorescence spectra measured after selective excitation of red chlorophylls show a decreased fluorescence of bulk chlorophylls around 685 nm, indicating that the thermal equilibration of the excitations between bulk and red chlorophylls is not complete prior to emission. Taking these results together, there seems to be almost no preference for Car S1 → red chlorophyll energy transfer, even though this could have been energetically favorable. We conclude that the small number of red chlorophylls is not sufficient to act as major acceptors for Car S1 energy. The fact that no increased red-chlorophyll emission is observed after two-photon excitation also provides a strong indication that no two-photon-allowed excitonic states can be formed from chlorophyll−chlorophyll or carotenoid−chlorophyll interactions. A comparison of the measured two-photon data with experimental and theoretical chlorophyll energies yields a fraction of ∼40% carotenoids that are potential Car S1 donors, neglecting any hot-state energy transfer. Spectral overlap calculations yield estimated vibrational ground-state Car S1 → Chl energy-transfer time constants of 3.1−4.4 ps.