Bernát Gábor, Morvaridi Fatemeh, Feyziyev Yashar, Styring Stenbjörn
Department of Biochemistry, Center for Chemistry and Chemical Engineering, Lund University, P.O. Box 124, S-221 00 Lund, Sweden.
Biochemistry. 2002 May 7;41(18):5830-43. doi: 10.1021/bi011691u.
We have investigated the pH dependence for each individual redox transition in the S-cycle of the oxygen evolving complex (OEC) of photosystem II by electron paramagnetic resonance (EPR) spectroscopy. In the experiments, OEC is advanced to the appropriate S-state at normal pH. Then, the pH is rapidly changed, and a new flash is given. The ability to advance to the next S-state in the cycle at different pHs is determined by measurements of the decrease or increase of characteristic EPR signals from the OEC in different S-states. In some cases the measured EPR signals are very small (this holds especially for the S0 ML signal at pH >7.5 and pH <4.8). Therefore, we refrain from providing error limits for the determined pK's. Our results indicate that the S1 --> S2 transition is independent of pH between 4.1 and 8.4. All other S-transitions are blocked at low pH. In the acidic region, the pK's for the inhibition of the S2 --> S3, the S3 --> [S4] --> S0, and the S0 --> S1 transitions are about 4.0, 4.5, and 4.7, respectively. The similarity of these pK values indicates that the inhibition of the steady-state oxygen evolution in the acidic range, which occurs with pK approximately 4.8, is a consequence of similar pH blocks in three of the redox steps involved in the oxygen evolution. In the alkaline region, we report a clear pH block in the S3 --> [S4] --> S0 transition with a pK of about 8.0. Our study also indicates the existence of a pH block at very high pH (pK approximately 9.4) in the S2 --> S3 transition. The S0 --> S1 transition is not affected, at least up to pH 9.0. This suggests that the inhibition of the steady-state oxygen evolution, which occurs with a pK of 8.0, is dominated by the inhibition of the S3 --> [S4] --> S0 transition. Our results are obtained in the presence of 5% methanol (v/v). However, it is unlikely that the determined pK's are affected by the presence of methanol since our results also show that the pH dependence of the steady-state oxygen evolution is not affected by methanol. The results in the alkaline region are in good agreement with a model, which suggests that the redox potential of Y(Z*)/Y(Z) is directly affected by high pH. At high pH the Y(Z*)/Y(Z) potential becomes lower than that of S2/S1 and S3/S2. The acidic block, with a pK of 4-5 in three S-transitions, implies that the inhibition mechanism is similar, and we suggest that it reflects protonation of a carboxylic side chain in the proton relay that expels protons from the OEC.
我们通过电子顺磁共振（EPR）光谱研究了光系统II放氧复合体（OEC）的S循环中各个氧化还原转变的pH依赖性。在实验中，OEC在正常pH下被推进到适当的S态。然后，迅速改变pH，并给予新的闪光。通过测量不同S态下OEC特征EPR信号的减少或增加，来确定在不同pH下推进到循环中下一个S态的能力。在某些情况下，测得的EPR信号非常小（在pH >7.5和pH <4.8时，S0 ML信号尤其如此）。因此，我们没有给出所确定的pK值的误差范围。我们的结果表明，S1→S2转变在4.1至8.4之间与pH无关。所有其他S转变在低pH下均受阻。在酸性区域，抑制S2→S3、S3→[S4]→S0和S0→S1转变的pK值分别约为4.0、4.5和4.7。这些pK值的相似性表明，在酸性范围内稳态放氧的抑制（pK约为4.8）是放氧过程中三个氧化还原步骤中类似pH阻断的结果。在碱性区域，我们报告了S3→[S4]→S0转变中存在明显的pH阻断，pK约为8.0。我们的研究还表明，在非常高的pH（pK约为9.4）下，S2→S3转变存在pH阻断。S0→S1转变至少在pH 9.0之前不受影响。这表明，pK为8.0时发生的稳态放氧抑制主要由S3→[S4]→S0转变的抑制主导。我们的结果是在5%甲醇（v/v）存在的情况下获得的。然而，所确定的pK值不太可能受到甲醇存在的影响，因为我们的结果还表明，稳态放氧的pH依赖性不受甲醇影响。碱性区域的结果与一个模型非常吻合，该模型表明Y(Z*)/Y(Z)的氧化还原电位直接受高pH影响。在高pH下，Y(Z*)/Y(Z)电位变得低于S2/S1和S3/S2的电位。在三个S转变中pK为4 - 5的酸性阻断意味着抑制机制相似，我们认为这反映了质子传递中一个羧基侧链的质子化，该质子化将质子从OEC中排出。
