The role of cytochrome b559 and tyrosine D in protection against photoinhibition during in vivo photoactivation of Photosystem II

Abstract

In vivo photoactivation of Photosystem II was studied in the FUD39 mutant strain of the green alga Chlamydomonas reinhardtii which lacks the 23 kDa protein subunit involved in water oxidation. Dark grown cells, devoid of oxygen evolution, were illuminated at 0.8 μE m −2 s −1 light intensity which promotes optimal activation of oxygen evolution, or at 17 μE m −2 s −1, where photoactivation compete with deleterious photodamage. The involvement of the two redox active cofactors tyrosine D and cytochrome b 559 during the photoactivation process, was investigated by EPR spectroscopy. Tyrosine D on the D 2 reaction center protein functions as auxiliary electron donor to the primary donor P 680 + during the first minutes of photoactivation at 0.8 μE m −2 s −1 (compare with Rova et al., Biochemistry, 37 (1998) 11039–11045.). Here we show that also cytochrome b 559 was rapidly oxidized during the first 10 min of photoactivation with a similar rate to tyrosine D. This implies that both cytochrome b 559 and tyrosine D may function as auxiliary electron donors to P 680 + and/or the oxidized tyrosine ⋅ Z on the D 1 protein, to avoid photoinhibition before successful photoactivation was accomplished. As the catalytic water-oxidation successively became activated, Tyrosine D remained oxidized while cytochrome b 559 became rereduced to the equilibrium level that was observed prior to photoactivation. At 17 μE m −2 s −1 light intensity, where photoinhibition competes significantly with photoactivation, tyrosine D was very rapidly completely oxidized, after which the amount of oxidized tyrosine D decreased due to photoinhibition. In contrast, cytochrome b 559 became reduced during the first 2 min of photoactivation at 17 μE m −2 s −1. After this, it was reoxidized, returning to the equilibrium level within 10 min. Thus, during in vivo photoactivation in high-light cytochrome b 559 serves two functions. Initially, it probably oxidizes the reduced primary acceptor pheophytin, thereby relieving the acceptor side of reductive pressure, and later on it serves as auxiliary electron donor, preventing donor-side photoinhibition.