Effect of Trypsin Treatment of Photosystem I Particles on the Electron Donation to P700

Abstract

Proteolysis of photosystem I particles had no effect on P700 oxidation but did inhibit the rate of P700+ reduction. The V,, values were decreased for both dichlorophenol and plastocyanin, but the Km values were unaffected indicating that trypsin treatment altered electron transfer rather than the binding of the donor to the photosystem I complex. The salt dependence of P700+ reduction was unaffected. The effects of P700+ reduction were the same for the preparations of different workers (Shiozawa, Alberte, Thornber 1974 Arch Biochem Biophys 165: 388; and Bengis, Nelson 1975 J Biol Chem 250: 2783).In both cases, the 70-kilodalton, chlorophyll-containing polypeptide was digested confirming its role in transferring electrons from plastocyanin to P700. The fact that the preparation of Shiozawa et at. lacks subunit (III) but still used plastocyanin as the electron donor rules out a role for this subunit as "the plastocyanin binding protein." Subunit III was also digested in the Bengis and Nelson preparation.Treatment with proteolytic enzymes has been used to study the structure and function of the chloroplast membrane. Experiments involving protease treatment of intact thylakoid membranes have shown that the pigment protein complexes were less susceptible to proteolytic attack than were the peripheral proteins (6,17,19,24). However, extensive proteolysis did lead to breakdown of the pigment protein complexes (6,14). In some cases, the fragmented polypeptides remained attached to the complex until the Chl molecules were removed (14, 16) indicating a structural role for the Chl molecules.We have extended these studies to include the effects of tryspin treatment on isolated PSI particles in order to identify the functions of the individual polypeptides in the complex. We have used two types of PSI preparations in these studies. One is the preparation of Shiozawa et al. (23) which contains Cyt b6 and Cytf in addition to a 70-kD polypeptide which contains Chl a and P700 (1). The second preparation is that of Bengis and Nelson (3,4) which contains (in addition to the 70-kD subunit), the secondary electron acceptors and a 20-kD subunit (subunit III) which has been implicated in the binding of plastocyanin (I 1).

Effects of Trypsin Treatment on the Photochemical Activity and Polypeptide Composition of the P700–ChlorophyllaProtein Complex

“…Essentially the same phenomena were observed after 24 hr of incubation with trypsin. Bhardwaj et al 6 ) have reported that proteolysis of isolated photosystem I particles had no effect on P700 oxidation but inhibited the rate of P700+ reduction by plastocyanin. Our results on trypsin treatment of the complex agree with the results of Bhardwaj et al 6 ) with respect to the effects of trypsin treatment of P700 photooxidation, but disagree with respect to the effects on the reduction rate of P700+ by plastocyanin.…”

“…6 ) Those results suggest that the complex retains its integrity to some extent even after the degradation of the polypeptides with tThis work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan.…”

The plastocyanin binding domain of photosystem I.

“…Haehnel, 1986) which can be partially reversed at high concentrations of magnesium ions (Ratajczak et al, 1988). This electron transfer to P700 in the absence of PsaF (Bhardwaj et al, 1982;Takabe et al, 1983) has been one of the reasons for suggesting that subunit PsaF may serve a different function. In some cases PsaF has been found to be associated with the peripheral light-harvesting complex (LHCI) of PSI (Anandan et al, 1989;Scheller and M0ller, 1990;Bassi et al, 1992).…”