Sétif P Q, Bottin H
C.E.A., Département de Biologie Cellulaire et Moléculaire, Gif sur Yvette, France.
Biochemistry. 1994 Jul 19;33(28):8495-504. doi: 10.1021/bi00194a014.
The kinetics of reduction of soluble ferredoxin by photosystem I (PSI), both purified from the cyanobacterium Synechocystis sp. PCC 6803, were investigated by flash-absorption spectroscopy between 460 and 600 nm. Most experiments were made with isolated monomeric PSI reaction centers prepared with the detergent beta-dodecyl maltoside. Analysis of absorption transients, in parallel at 480 and 580 nm and under several conditions, shows the existence of three different first-order components in the presence of ferredoxin (t1/2 approximately 500 ns, 20 microseconds, and 100 microseconds). A second-order phase of ferredoxin reduction is also present [k = (2-5) x 10(8) s-1 at pH 8 and at moderate ionic strength]. Similar first-order kinetic components were found with membranes from Synechocystis, with dissolved crystals of trimeric PSI reaction centers from Synechococcus, and also when ferredoxin from Synechocystis is replaced by ferredoxin from Chlamydomonas reinhardtii. The three first-order phases exhibit similar, though not identical, spectra which are consistent with electron transfer from the [4Fe-4S] centers of PSI to the [2Fe-2S] center of ferredoxin and are all attributed to reduction of ferredoxin bound to PSI. At pH 8 and at moderate ionic strength, the dissociation constants associated with each of these components are also similar, with a global value varying between 0.2 and 0.8 microM in different cyanobacterial preparations. The presence of three exponential components is discussed assuming homogeneity of the two partners and using the estimated values for the shortest possible distance of approach of soluble ferredoxin from the different iron-sulfur centers of PSI. It is concluded that the 500-ns phase corresponds to electron transfer from either FA- or FB-, the terminal iron-sulfur acceptors of PSI, to ferredoxin and that the immediate electron donor to ferredoxin is reduced within less than 500 ns. The presence of at least two different types of PSI-ferredoxin complex, all competent in electron transfer, is also deduced from the kinetic behavior.
利用460至600纳米波长范围内的闪光吸收光谱，研究了从集胞藻属蓝藻PCC 6803中纯化得到的光系统I（PSI）还原可溶性铁氧化还原蛋白的动力学。大多数实验是使用由去污剂β - 十二烷基麦芽糖苷制备的分离单体PSI反应中心进行的。在480和580纳米波长并行且在几种条件下对吸收瞬态进行分析，结果表明在存在铁氧化还原蛋白的情况下存在三种不同的一级成分（半衰期约为500纳秒、20微秒和100微秒）。铁氧化还原蛋白还原还存在一个二级相[k = (2 - 5)×10⁸ s⁻¹，在pH 8和中等离子强度下]。在集胞藻的膜、来自聚球藻的三聚体PSI反应中心的溶解晶体中，以及当集胞藻的铁氧化还原蛋白被莱茵衣藻的铁氧化还原蛋白替代时，都发现了类似的一级动力学成分。这三个一级相表现出相似但不完全相同的光谱，这与电子从PSI的[4Fe - 4S]中心转移到铁氧化还原蛋白的[2Fe - 2S]中心一致，并且都归因于与PSI结合的铁氧化还原蛋白的还原。在pH 8和中等离子强度下，与这些成分各自相关的解离常数也相似，在不同蓝藻制剂中全局值在0.2至0.8微摩尔之间变化。假设两个伙伴具有同质性，并使用可溶性铁氧化还原蛋白从PSI不同铁硫中心接近的最短可能距离的估计值，对三个指数成分的存在进行了讨论。得出的结论是，500纳秒相对应于电子从PSI的末端铁硫受体FA - 或FB - 转移到铁氧化还原蛋白，并且铁氧化还原蛋白的直接电子供体在不到500纳秒内被还原。从动力学行为还推断出至少存在两种不同类型的PSI - 铁氧化还原蛋白复合物，它们都能够进行电子转移。
