Interaction of ferredoxin:NADP+ oxidoreductase with phycobilisomes and phycobilisome substructures of the cyanobacterium Synechococcus sp strain PCC 7002
Interaction of ferredoxin:NADP+ oxidoreductase with phycobilisomes and phycobilisome substructures of the cyanobacterium Synechococcus sp strain PCC 7002
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DOI：
10.1021/bi0346998
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发表时间：
2003-12-02
期刊：
影响因子：
2.9
通讯作者：
Bryant, DA
中科院分区：
文献类型：
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作者：
Gómez-Lojero, C;Pérez-Gómez, B;Bryant, DA
来自Synechococcus sp. PCC 7002的酶铁氧还蛋白-NADP（+）氧化还原酶（FNR）具有包含三个结构域（FNR-3D）的延伸结构（Schluchter，W. M.，和Bryant，D. A.（1992）Biochemistry 31，3092-3102）。来自野生型细胞的藻胆体（PBS）制备物含有1.0至1.6个FNR-3D分子/PBS，平均值为1.3个FNR/PBS。当加入外源性重组FNR-3D（rFNR-3D）时，最多两个FNR-3D分子可以通过酶的N-末端CpcD样结构域与野生型PBS特异性结合。为了将酶定位在PBS内，进一步研究了PBS及其亚结构与rFNR-3D的相互作用。rFNR-3D对藻蓝蛋白（PC）六聚体的结合亲和力高于其对不含接头蛋白的PC六聚体的亲和力，其中藻蓝蛋白（PC）六聚体含有来源于CpcG（L-RC（27）接头多肽）的22-kDa蛋白水解片段。来自cpcD 3突变体的PBS，其缺乏9-kDa的PC相关的杆状接头，每个PBS掺入多达6个rFNR-3D分子。cpcC突变体的PBS，其具有含有单个PC六聚体的外周杆，每个PBS也掺入多达6个rFNR-3D分子。直接竞争结合实验表明，来自cpcD 3突变体的PBS比来自cpcC突变体的PBS结合更多的酶。这些观察结果支持这样的假设，即酶优先结合到PBS的外周杆的远端。这些数据还表明，PC复合物对FNR-3D的相对亲和力顺序如下：（α（PC）β（PC））（6）-L-R（33）>（α（PC x）β（PC））（6）-L-RC（27），>（α（PC）β（PC））（6）。这些数据表明，在PBS的组装过程中，FNR-3D可以根据其对不同PC亚复合物的相对结合亲和力被置换到外周。因此，FNR-3D不会干扰PBS外周杆中PC的光吸收和能量转移特性。这种本地化的PBS内的FNR与其功能在蓝藻的影响进行了讨论。
The enzyme ferredoxin-NADP(+) oxidoreductase (FNR) from Synechococcus sp. PCC 7002 has an extended structure comprising three domains (FNR-3D) (Schluchter, W. M., and Bryant, D. A. (1992) Biochemistry 31, 3092-3102). Phycobilisome (PBS) preparations from wild-type cells contained from 1.0 to 1.6 molecules of FNR-3D per PBS, with an average value of 1.3 FNR per PBS. A maximum of two FNR-3D molecules could be specifically bound to wild-type PBS via the N-terminal, CpcD-like domain of the enzyme when exogenous recombinant FNR-3D (rFNR-3D) was added. To localize the enzyme within the PBS, the interaction of PBS and their substructures with rFNR-3D was further investigated. The binding affinity of rFNR-3D for phycocyanin (PC) hexamers, which contained a 22-kDa proteolytic fragment derived from CpcG, the L-RC(27) linker polypeptide, was higher than its affinity for PC hexamers containing no linker protein. PBS from a cpcD3 mutant, which lacks the 9-kDa, PC-associated rod linker, incorporated up to six rFNR-3D molecules per PBS. PBS of a cpcC mutant, which has peripheral rods that contain single PC hexamers, also incorporated up to six rFNR-3D molecules per PBS. Direct competition binding experiments showed that PBS from the cpcD3 mutant bound more enzyme than PBS from the cpcC mutant. These observations support the hypothesis that the enzyme binds preferentially to the distal ends of the peripheral rods of the PBS. These data also show that the relative affinity order of the PC complexes for FNR-3D is as follows: (alpha(PC)beta(PC))(6)-L-R(33) > (alpha(PCx)beta(PC))(6)-L-RC(27), > (alpha(PC)beta(PC))(6). The data suggest that, during the assembly of the PBS, FNR-3D could be displaced to the periphery according to its relative binding affinity for different PC subcomplexes. Thus, FNR-3D would not interfere with the light absorption and energy transfer properties of PC in the peripheral rods of the PBS. The implications of this localization of FNR within the PBS with respect to its function in cyanobacteria are discussed.