FLUORESCENCE QUENCHING BY CHLOROPHYLL CATIONS IN PHOTOSYSTEM-II

Although fluorescence is widely used to study photosynthetic systems, the mechanisms that affect the fluorescence in photosystem II (PSII) a re not completely understood. The aim of this study is to define the l ow-temperature steady-state fluorescence quenching of redox-active cen ters that function on the electron donor side of PSII. The redox state s of the electron donors and accepters were systematically varied by u sing a combination of pretreatments and illumination to produce and tr ap, at low temperature, a specific charge-separated state. Electron pa ramagnetic resonance spectroscopy and fluorescence intensity measureme nts were carried out on the same samples to obtain a correlation betwe en the redox state and the fluorescence. It was found that illuminatio n of PSII at temperatures between 85 and 260 K induced a fluorescence quenching state in two phases. At 85 K, where the fast phase was most prominent, only one electron-transfer pathway is active on the donor s ide of PSII. This pathway involves electron donation to the primary el ectron donor in PSII, P680, from cytochrome b(559) and a redox-active chlorophyll molecule, Chl(Z). Oxidized Chl(Z) was found to be a potent quencher of chlorophyll fluorescence with 15% of oxidized Chl(Z) suff icient to quench 70% of the fluorescence intensity. This implies that neighboring PSII reaction centers are energetically connected, allowin g oxidized Chl(Z) in a few centers to quench most of the fluorescence. The presence of a well-defined quencher in PSII may make it possible to study the connectivity between antenna systems in different sample preparations. The other redox-active components on the donor side of P SII studied were the O-2-evolving complex, the redox-active tyrosines (Y-Z and Y-D), and cytochrome b(559). No significant changes in fluore scence intensity could be attributed to changes in the redox state of these components. The fast phase of fluorescence quenching is attribut ed to the rapid photooxidation of Chl(Z), and the slow phase is attrib uted to multiple turnovers providing for further oxidation of Chl(Z) a nd irreversible photoinhibition. Significant photoinhibition only occu rred at Chl concentrations below 0.7 mg/mL and above 150 K. The revers ible oxidation of Chl(Z) in intact systems may function as a photoprot ection mechanism under high-light conditions and account for a portion of the nonphotochemical fluorescence quenching.