Biochemical and structural characterization of Pseudomonas aeruginosa Bfd and FPR: ferredoxin NADP+ reductase and not ferredoxin is the redox partner of heme oxygenase under iron-starvation conditions.
Biochemical and structural characterization of Pseudomonas aeruginosa Bfd and FPR: ferredoxin NADP+ reductase and not ferredoxin is the redox partner of heme oxygenase under iron-starvation conditions.
复制标题
铜绿假单胞菌 Bfd 和 FPR 的生化和结构特征：铁氧还蛋白 NADP 还原酶而非铁氧还蛋白是缺铁条件下血红素加氧酶的氧化还原伴侣。
DOI：
10.1021/bi7013135
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发表时间：
2007
期刊：
影响因子：
2.9
通讯作者：
Rivera,Mario
中科院分区：
文献类型：
--
作者：
Wang,An;Zeng,Yuhong;Han,Huijong;Weeratunga,Saroja;Morgan,BaileyN;Moënne-Loccoz,Pierre;Schönbrunn,Ernst;Rivera,Mario
在118个基因中，铜绿假单胞菌在铁饥饿反应中上调[Ochsner，U.一、Wilderman，P. J.，Vasil，A.一、和Vasil，M. L.等人（2002）Mol. Microbiol.45，1277−1287]，我们专注于编码电子转移蛋白的两个基因的产物，作为鉴定在低铁胁迫条件下表达的血红素加氧酶（pa-HO）的氧化还原伴侣的手段。生物化学和光谱学研究表明，bfd基因编码一个含有[2Fe-2S]2+/+中心的73个氨基酸的蛋白质（pa-Bfd），而fpr基因编码一个258个残基的NADPH依赖性铁氧还蛋白还原酶（pa-FPR），它利用FAD作为辅因子。在体外重建的pa-HO催化活性与新表征的蛋白质导致令人惊讶的观察，pa-FPR有效地支持pa-HO的催化循环，而不需要铁氧还蛋白。相比之下，电子转移从pa-Bfd topa-HO是缓慢的，这强烈反对的可能性，即所需的7个电子由pa-HO降解胆绿素是从NADPH topa-HO铁氧还蛋白（Bfd）依赖的方式转移。考虑到pa-HO的功能是从铁饥饿条件下获得的外源血红素中释放铁，使用黄素酶而不是含铁硫中心的蛋白质来支持血红素降解是对细胞资源的有效利用。pa-FPR的晶体结构（1.6 μ m分辨率）表明，它的折叠与铁氧还蛋白还原酶超家族的折叠相当，与棕色固氮菌（Azotobacter vinelandii）FPR和大肠杆菌（Escherichia coli）铁氧还蛋白还原酶的结构最相似。后两种酶分别与不同的氧化还原伙伴铁氧还蛋白和黄素氧还蛋白相互作用。因此，本文报道的发现扩展了由pa-FPR折叠识别的氧化还原配偶体的范围，以包括血红素加氧酶（pa-HO）。
Among the 118 genes upregulated byPseudomonas aeruginosain response to iron starvation [Ochsner, U. A., Wilderman, P. J., Vasil, A. I., and Vasil, M. L. (2002)Mol. Microbiol.45, 1277−1287], we focused on the products of the two genes encoding electron transfer proteins, as a means of identifying the redox partners of the heme oxygenase (pa-HO) expressed under low-iron stress conditions. Biochemical and spectroscopic investigations demonstrated that thebfdgene encodes a 73-amino acid protein (pa-Bfd) that incorporates a [2Fe-2S]2+/+center, whereas thefprgene encodes a 258-residue NADPH-dependent ferredoxin reductase (pa-FPR) that utilizes FAD as a cofactor. In vitro reconstitution ofpa-HO catalytic activity with the newly characterized proteins led to the surprising observation thatpa-FPR efficiently supports the catalytic cycle ofpa-HO, without the need of a ferredoxin. In comparison, electron transfer frompa-Bfd topa-HO is sluggish, which strongly argues against the possibility that the seven electrons needed bypa-HO to degrade biliverdin are transferred from NADPH topa-HO in a ferredoxin (Bfd)-dependent manner. Given thatpa-HO functions to release iron from exogenous heme acquired under iron-starvation conditions, the use of a flavoenzyme rather than an iron−sulfur center-containing protein to support heme degradation is an efficient use of resources in the cell. The crystal structure ofpa-FPR (1.6 Å resolution) showed that its fold is comparable that of the superfamily of ferredoxin reductases and most similar to the structure ofAzotobacter vinelandiiFPR andEscherichia coliflavodoxin reductase. The latter two enzymes interact with distinct redox partners, a ferredoxin and a flavodoxin, respectively. Hence, findings reported herein extend the range of redox partners recognized by the fold ofpa-FPR to include a heme oxygenase (pa-HO).