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-Sta

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• 作者：An Wang ; Yuhong Zeng ; Huijong Han ; Saroja Weeratunga ; Bailey N. Morgan ; Pierre Mo&euml ; nne-Loccoz ; Ernst Schö ; nbrunn ; Mario Rivera
• 刊名：Biochemistry
• 出版年：2007
• 出版时间：October 30, 2007
• 年：2007
• 卷：46
• 期：43
• 页码：12198 - 12211
• 全文大小：512K
• 年卷期：v.46,no.43(October 30, 2007)
• ISSN：1520-4995

文摘

Among the 118 genes upregulated by Pseudomonas aeruginosa in 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 identifyingthe redox partners of the heme oxygenase (pa-HO) expressed under low-iron stress conditions. Biochemicaland spectroscopic investigations demonstrated that the bfd gene encodes a 73-amino acid protein (pa-Bfd) that incorporates a [2Fe-2S]^2+/+ center, whereas the fpr gene encodes a 258-residue NADPH-dependentferredoxin reductase (pa-FPR) that utilizes FAD as a cofactor. In vitro reconstitution of pa-HO catalyticactivity with the newly characterized proteins led to the surprising observation that pa-FPR efficientlysupports the catalytic cycle of pa-HO, without the need of a ferredoxin. In comparison, electron transferfrom pa-Bfd to pa-HO is sluggish, which strongly argues against the possibility that the seven electronsneeded by pa-HO to degrade biliverdin are transferred from NADPH to pa-HO in a ferredoxin (Bfd)-dependent manner. Given that pa-HO functions to release iron from exogenous heme acquired underiron-starvation conditions, the use of a flavoenzyme rather than an iron-sulfur center-containing proteinto support heme degradation is an efficient use of resources in the cell. The crystal structure of pa-FPR(1.6 Å resolution) showed that its fold is comparable that of the superfamily of ferredoxin reductases andmost similar to the structure of Azotobacter vinelandii FPR and Escherichia coli flavodoxin 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 of pa-FPR toinclude a heme oxygenase (pa-HO).