Boussac A, Rutherford A W, Styring S
Département de Biologie, CEN Saclay, Gif-sur-Yvette, France.
Biochemistry. 1990 Jan 9;29(1):24-32. doi: 10.1021/bi00453a003.
The effects of NH3 on the oxygen evolving enzyme have been investigated with EPR and steady-state O2 evolution. The following results were obtained. At low light intensity O2 evolution occurs in all centers even though ammonia is bound. This binding occurs in the S2 state and results in a modification of the multiline signal as reported earlier. However, the oscillations with flash number of the amplitude of the EPR signal are virtually unaffected, indicating that NH3 binding does not prevent S-state advancement. Inhibition of O2 evolution by NH3 measured at light intensities that are nearly saturating for untreated photosystem II is interpreted as being due to a slow down in the rate of S-state cycling. At very high light intensities NH3 is not able to inhibit oxygen evolution presumably because NH3 binding is S state dependent and the susceptible S state (S2) is turned over too quickly. NH3 binding resulting in the modified multiline signal does not occur in S1. When S1 is formed from fully NH3 modified S2 by deactivation or by three further flashes, the S1 state does not have NH3 bound. NH3 thus dissociates easily from S1. Earlier reports of NH3 binding in S1 may be explained by the observation that NH3 binding can occur upon incubation of samples in S2 at temperatures as low as 198 K. Evidence is obtained for an NH3 binding occurring slowly (30 s) in S3. This binding results in a block in S-state advancement as suggested earlier [Velthuys, B. R. (1975) Thesis, University of Leiden]. The results are interpreted in two possible models: (1) NH3 binding in S2 occurs in a substrate site, but it is rapidly exchanged by water upon S4 formation. (2) NH3 binding in S2 is not in a substrate site but instead in a structural site and remains bound while water is oxidized. Inherent in this model is that other NH3 binding sites, i.e., the Cl- site, and the slow NH3 binding site in S3 could be the true substrate sites. Some mechanistic implications are discussed.
利用电子顺磁共振（EPR）和稳态放氧对氨（NH₃）对放氧酶的影响进行了研究，得到以下结果。在低光强下，即使氨已结合，所有中心仍会发生放氧。这种结合发生在S₂状态，并如先前报道的那样导致多线信号的改变。然而，EPR信号幅度随闪光次数的振荡几乎不受影响，这表明NH₃结合并不妨碍S态的推进。在几乎使未处理的光系统II达到饱和的光强下测量，NH₃对放氧的抑制作用被解释为是由于S态循环速率减慢所致。在非常高的光强下，NH₃无法抑制放氧，大概是因为NH₃结合依赖于S态，而易感的S态（S₂）转换太快。在S₁中不会发生导致多线信号改变的NH₃结合。当通过失活或再经过三次闪光从完全被NH₃修饰的S₂形成S₁时，S₁状态没有结合NH₃。因此，NH₃很容易从S₁解离。先前关于S₁中NH₃结合的报道可能可以通过以下观察结果来解释：在低至198K的温度下，将样品在S₂中孵育时会发生NH₃结合。有证据表明在S₃中会缓慢（30秒）发生NH₃结合。如先前[韦尔图斯，B.R.（1975年）论文，莱顿大学]所指出的，这种结合导致S态推进受阻。这些结果在两种可能的模型中得到解释：（1）S₂中的NH₃结合发生在底物位点，但在形成S₄时会迅速被水交换。（2）S₂中的NH₃结合不在底物位点，而是在结构位点，并且在水被氧化时仍然结合。该模型的内在含义是，其他NH₃结合位点，即Cl⁻位点和S₃中的缓慢NH₃结合位点可能是真正的底物位点。讨论了一些机制方面的影响。
