The role of cytochrome b(559) and tyrosine(D) in protection against photoinhibition during in vivo photoactivation of photosystem II

In vivo photoactivation of Photosystem II was studied in the FUD39 mutant s train of the green alga Chlamydomonas reinhardtii which lacks the 23 kDa pr otein subunit involved in water oxidation. Dark grown cells, devoid of oxyg en evolution, were illuminated at 0.8 mu E m(-2) s(-1) light intensity whic h promotes optimal activation of oxygen evolution, or at 17 mu E m(-2) s(-1 ), where photoactivation compete with deleterious photodamage. The involvem ent of the two redox active cofactors tyrosine(D) and cytochrome b(559) dur ing the photoactivation process, was investigated by EPR spectroscopy. Tyro sine(D) on the D-2 reaction center protein functions as auxiliary electron donor to the primary donor P-680(+) during the first minutes of photoactiva tion at 0.8 mu E m(-2) s(-1) (compare with Rova et al., Biochemistry, 37 (1 998) 11039-11045.). Here we show that also cytochrome b(559) was rapidly ox idized during the first 10 min of photoactivation with a similar rate to ty rosine(D). This implies that both cytochrome b(559) and tyrosine(D), may fu nction as auxiliary electron donors to P-680(+) and/or the oxidized tyrosin e; on the D-1 protein, to avoid photoinhibition before successful photoacti vation was accomplished. As the catalytic water-oxidation successively beca me activated, Tyrosine(D) remained oxidized while cytochrome b(559) became rereduced to the equilibrium level that was observed prior to photoactivati on. At 17 mu E 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 mu E m(-2) s(-1). After this, it was r eoxidized, returning to the equilibrium level within 10 min. Thus, during i n 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. (C) 1999 Elsevier Science B.V. All rights reserved.