A New Pathway of Photoinactivation of Photosystem II. Irreversible Photoreduction of Pheophytin Causes Loss of Photochemical Activity of Isolated D1-D2-Cytochrome b₅₅₉ Complex

O. V. Nikitishena^*, T. N. Smolova, R. A. Khatypov, A. Ja. Shkuropatov, and V. V. Klimov

Institute of Fundamental Problems of Biology, Russian Academy of Sciences, Pushchino, Moscow Region, 142290 Russia; fax: (27) 79-0532; E-mail: lwomain@issp.serpukhov.su

^* To whom correspondence should be addressed.

Received March 26, 2001; Revision received June 25, 2001

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A new pathway of photoinactivation of photosystem II (PS II) connected with irreversible photoaccumulation of reduced pheophytin (Ph) in isolated D1-D2-cytochrome b₅₅₉ complexes of reaction center (RC) of PS II was discovered. The inhibitory effects of white light illumination on photochemical activity of D1-D2-cytochrome b₅₅₉ complexes of RCs of photosystem II, isolated from pea chloroplasts, have been compared under anaerobic conditions in the absence and in the presence of sodium dithionite, electron transfer from which to the oxidized primary electron donor P680⁺ results in the photoaccumulation of anion-radical of the primary electron acceptor, Ph^-*. In both cases, prolonged illumination (1-5 min, 120 W/m²) led to a pronounced loss of the photochemical activity as it was monitored by measuring the amplitude of the reversible photoinduced absorbance changes at 682 nm related to the photoreduction of Ph. The extent of the photoinactivation depended on the illumination time and pH of the medium. At pH 8.0, the presence of dithionite during photoinactivation brought about a protective effect compared to that in a control sample. In contrast, lowering pH to 6.0 increased the sensitivity to photoinactivation in the dithionite-containing samples. For 5 min irradiation, the photochemical activity in the absence and in the presence of dithionite decreased by 35 and 72%, respectively (this was accompanied by an irreversible bleaching of the pheophytin Q_x absorption band at 542 nm). Degradation of the D1 and D2 proteins was not observed under these conditions. A subsequent addition of an electron acceptor, potassium ferricyanide, to the illuminated samples restored neither the amplitude of the signal at 682 nm nor absorption at 542 nm. It is suggested that at pH . 7.0 the photoaccumulated Ph^-* is irreversibly converted into a secondary, most probably protonated form, that does not lead to destruction of the RCs but prevents the photoformation of the primary radical pair [P680⁺Ph^-*]. A possible application of this effect to photoinactivation of PS II in vivo is discussed.

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KEY WORDS: photosystem II, reaction center, pheophytin, photoinactivation

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