Figure 1.
Light-induced TEMPONE EPR spectra measured in Tris-treated PSII membranes.
[A] Tris-treated PSII membranes (200 µg Chl ml−1) were illuminated with white light (1000 µmol m−2 s−1) in the presence of 50 mM TMPD and 40 mM MES-NaOH (pH 6.5) for the time period as indicated in figure. [B] Time profile of TEMPONE EPR signal measured in Tris-treated PSII membranes under light illumination. The data represent the mean value (±SD) of at least three experiments.
Figure 2.
Light-induced POBN-R adduct EPR spectra measured in Tris-treated PSII membranes.
[A] Tris-treated PSII membranes (200 µg Chl ml−1) were illuminated with white light (1000 µmol m−2 s−1) in the presence of 50 mM POBN and 40 mM MES-NaOH (pH 6.5) for the time period as indicated in figure. [B] Time profile of POBN-R adduct EPR signal measured in Tris-treated PSII membranes under light illumination. The data represent the mean value (±SD) of at least three experiments.
Figure 3.
Effect of pH on TEMPONE EPR spectra measured in Tris-treated PSII membranes.
[A] Tris-treated PSII membranes (200 µg Chl ml−1) were illuminated with white light (1000 µmol m−2 s−1) for 30 min at different pH as indicated in figure. The measurements were performed in 40 mM MES buffer (pH 6) or 40 mM HEPES buffer (pH 7) or 40 mM TRIS buffer (pH 8) or 40 mM CAPSO buffer (pH 9) or 40 mM CASP buffer (pH 10). [B] pH profile of TEMPONE EPR signal measured in Tris-treated PSII membranes under light illumination for 30 min. The intensity of EPR signal was evaluated as the relative height of central peak of the first derivative of the EPR absorption spectrum. The data represent the mean value (±SD) of at least three experiments.
Figure 4.
Effect of pH on POBN-R adduct EPR spectra measured in Tris-treated PSII membranes.
[A] Tris-treated PSII membranes (200 µg Chl ml−1) were illuminated with white light (1000 µmol m−2 s−1) for 30 min at different pH as indicated in figure. The measurements were performed in 40 mM MES buffer (pH 6) or 40 mM HEPES buffer (pH 7) or 40 mM TRIS buffer (pH 8) or 40 mM CAPSO buffer (pH 9) or 40 mM CASP buffer (pH 10). [B] pH profile of POBN-R adduct EPR signal measured in Tris-treated PSII membranes under light illumination for 30 min. The intensity of EPR signal was evaluated as the relative height of central peak of the first derivative of the EPR absorption spectrum. The data represent the mean value (±SD) of at least three experiments.
Figure 5.
Effect of propyl gallate on TEMPONE and EMPO-R adduct EPR spectra measured in Tris-treated PSII membranes.
[A] Tris-treated PSII membranes (200 µg Chl ml−1) were illuminated with white light (1000 µmol m−2 s−1) for 30 min in the absence (Light) and the presence (Light+PG) of 5 mM propyl gallate. Other experimental conditions were the same as described in Figure 1. [B] Tris-treated PSII membranes (200 µg Chl ml−1) were illuminated with white light (1000 µmol m−2 s−1) for 30 min in 40 mM MES-NaOH (pH 6.5), then added spin trap EMPO and measured the spectra without further illumination (dark). Due to the instability of EMPO-R adduct for a longer period illumination, Tris-treated PSII membranes were first illuminated 30 min in the absence of spin trap and then illuminated for additional 2 min in the presence of spin trap EMPO (Light). Tris-treated PSII membranes were first illuminated 30 min in the in the presence of 5 mM propyl gallate and then illuminated for additional 2 min by adding spin trap EMPO (Light+PG).
Figure 6.
Proposed mechanism for the formation of 1O2 in the donor side photoinhibition of PSII.
Carbon centered radical (R•) formed by oxidation of lipids and proteins react with molecular oxygen to form peroxyl radical (ROO•). Two peroxyl radicals react with the each other to form linear tetraoxide (ROOOOR) known to decompose to singlet oxygen (1O2), carbonyl (RO) and alcohols (ROH) via the Russell mechanism.