Under certain conditions the isolated reaction centre (RC) of photosystem 2 (PS 2) ís highly vulnerahle to photoinduced damage. With no added secondary electron acceptors or donors tíďs damage is due to singlet oxygen generated by the P680 triplet. This triplet is formed by recombination of the radical pair PóSO+Pheo' and Ihe photoinduced damage only occurs under aerobic conditions. When an electron acceptor is present, the degradation of pigments and the Dl and D2 proteins is due to formation of P680+ and secondary oxidation processes. Under the latter but not the former condition, characteristic pattems of degradation firagments of the Dl and D2 proteins are observed. In particular 24 and 17 kDa breakdown firagments of Dl are obtained while the D2 protein yields firagments having molecular masses of 29 and 21 kDa. Experiments involving the use of antibodies, radiophosphate and speciííc proteolytic digestion indicate that all four firagments contain the C-terminal portions of their mature proteins. These findings indicate that the proteolytic cleavage sites are positioned on the lumenal side of the membrane, particularly in the region spanning transmembrane helices I and II. Related studies on the 24 lď)a Dl protein fragment generated in vivo using Synechocystis sp. PCC 6803 by photoinhibitory treatment give the same conclusion for this firagment. Such a conclusion seems to contrast with the previous suggestion that the initial cleavage of Dl protein associated with its degradatíon and tumover occurs on the outer side of membrane in the region spanning transmembrane helices IV and V.
A double ínductíon mechanism of Dl protein degradatíon in isolated photosystem 2 (PS2) core complexes and reaction centres is described, showing the existence of two potentíal sites for primáty cleavage. Donor side inhibition conditi- o n s (presence of electron acceptors but no electron donors and pH 8.0) trigger the hydrolysis of the Dl protein between the putatíve helices 1 and II on the lumenal side of the thylakoid membrane. This results in the generation of a C-terminal 24 kDa fragment. However, when the donor-side is actíve (presence of electron donors but no electron acceptors and pH 6.0, acceptor side inhibition conditions) both preparations are able to produce a N-terminal 23 kDa fragment, indicating cleavage between helices IV and V, on the stromal side of the membrane.