Stein R R, Castellvi A L, Bogacz J P, Wraight C A
J Cell Biochem. 1984;24(3):243-59. doi: 10.1002/jcb.240240306.
A select group of herbicides that inhibit photosystem II also act at the acceptor side of the reaction center (RC) from the photosynthetic bacterium Rhodopseudomonas sphaeroides, with much the same relative specificity as in plants. These include the triazines and some phenolic compounds. The proposal that herbicides inhibit the electron transfer from the primary quinone (QA) to the secondary quinone (QB) by competing for the secondary quinone binding site--the B-site--[5], is tested here with terbutryn, the most potent of the triazines. Competition between terbutryn and ubiquinone (Q-10) was observed using the kinetics of the back-reaction as a measure of inhibition. The model includes binding equilibria before and after flash activation. The binding constants for the preflash (dark) equilibria, for reaction centers in 0.14% lauryl dimethylamine-N-oxide (LDAO), were KDi = 0.8 microM terbutryn, KDq = 2 microM Q-10; both are detergent-concentration dependent. After flash activation, binding equilibrium is not fully restored on the time scale of the back-reaction because terbutryn unbinds slowly. This gives rise to biphasic decay kinetics from which koff for terbutryn was estimated to be 3 sec-1. Titrations of the rate of the slow back reaction indicated that the post-flash equilibrium is less sensitive to inhibitor, in a manner that is independent of the much stronger binding of the semiquinone, Q-B, and indicative of a direct effect of the redox state of QA on the affinity of the B-site for ligands. However, the effects on KLi and KDq could not be separated: either KLi greater than KDi or KLq less than KDq. Some triazine-resistant mutants have been isolated and are described. All appear to be herbicide binding site mutants. Whole cells and photosynthetic membrane vesicles (chromatophores) exhibit a 10-50-fold increase in resistance to triazines due, in large part, to an increase in the rate of unbinding (koff). The modifications of the binding site appear to diminish the affinity of the B-site for ubiquinone as well as terbutryn. It is concluded that bacterial RCs are a useful model for the study of herbicide activity and specificity.
一组能够抑制光系统II的除草剂也作用于光合细菌球形红假单胞菌反应中心（RC）的受体侧，其相对特异性与在植物中大致相同。这些除草剂包括三嗪类和一些酚类化合物。有人提出除草剂通过竞争次级醌结合位点（B位点）来抑制从初级醌（QA）到次级醌（QB）的电子转移[5]，本文用最有效的三嗪类除草剂特丁净对此进行了验证。利用反向反应动力学来衡量抑制作用，观察了特丁净与泛醌（Q-10）之间的竞争。该模型包括闪光激活前后的结合平衡。在0.14%月桂基二甲基氧化胺（LDAO）中，反应中心的预闪光（黑暗）平衡的结合常数为：KDi = 0.8 μM特丁净，KDq = 2 μM Q-10；两者均依赖于去污剂浓度。闪光激活后，由于特丁净解离缓慢，在反向反应的时间尺度上结合平衡未完全恢复。这导致了双相衰减动力学，据此估计特丁净的解离常数koff为3 s-1。对缓慢反向反应速率的滴定表明，闪光后平衡对抑制剂的敏感性较低，其方式与半醌Q-B更强的结合无关，这表明QA的氧化还原状态对B位点与配体亲和力有直接影响。然而，对KLi和KDq的影响无法区分：要么KLi大于KDi，要么KLq小于KDq。已分离出一些抗三嗪类除草剂的突变体并进行了描述。所有这些突变体似乎都是除草剂结合位点突变体。完整细胞和光合膜囊泡（载色体）对三嗪类除草剂的抗性增加了10 - 50倍，这在很大程度上是由于解离速率（koff）的增加。结合位点的修饰似乎降低了B位点对泛醌以及特丁净的亲和力。结论是细菌反应中心是研究除草剂活性和特异性的有用模型。
