In an earlier study it was demonstrated that multiple electron flow through Photosystem II (PS II, including apparently the S-states, can be sustained without detectable O2 evolution upon treatment of samples with the lipid analog compound lauroylcholine chloride (LCC) (Wydrzynski et al. (1985) Biochim. Biophys. Acta 809, 125-136). The question remained, however, as to what is the source of the electrons under this condition. In this study we extend our observations of the LCC effect and show that a peroxide-type intermediate is transiently formed on the donor side of PS II. Upon using the luminol/peroxidase method to detect peroxide, a new chemiluminescent signal (S(D)) appears after the illumination of LCC-treated samples. This new signal is kinetically distinct (t(max) = 2-3 s) from the chemiluminescent signal (S(A), t(max) almost-equal-to 0.2 s) ascribable to the free H2O2 formed by the reduction of O2 on the acceptor side of PS II (Ananyev and Klimov (1989) Biochemistry (USSR) 54, 1587-1597), which appears in both control and LCC-treated samples. Both S(A) and S(D) are sensitive to the PS II inhibitor DCMU to about the same extent. In the presence of low levels of exogenously added catalase (about 50-mu-g/ml), S(A) can be completely eliminated but not S(D), which disappears only at higher catalase concentrations. It is suggested that the peroxide which gives rise to S(D) is initially sequestered within the sample. After the complete removal of the functional manganese by a TEMED extraction procedure, S(D) is lost but not S(A), with or without the addition of the artificial electron donor, hydroxylamine. In a sequence of brief (1-mu-s) light flashes, the progressive differences in S(D) amplitudes exhibit periodic behavior, beginning on the first flash, but without a consistent pattern. As the flash sequence advances, a consumption of the peroxide appears to take place. The results are consistent with the hypothesis that in LCC-treated samples, the water-splitting catalytic complex is perturbed in such a way so that the substrate water molecules are not fully oxidized, to O2.
