Insight into the radical mechanism of phycocyanobilin-ferredoxin oxidoreductase (PcyA) revealed by X-ray crystallography and biochemical measurements.
The X-ray crystal structure of the substrate-free form of phycocyanobilin (PCB)-ferredoxin oxidoreductase (PcyA; EC 1.3.7.5) from the cyanobacterium Nostoc sp. PCC7120 has been solved at 2.5 A resolution. A comparative analysis of this structure with those recently reported for substrate-bound and substrate-free forms of PcyA from the cyanobacterium Synechocystis sp. PCC6803 (Hagiwara et al. (2006) Proc. Natl. Acad. Sci. U.S.A. 103, 27-32; Hagiwara et al. (2006) FEBS Lett. 580, 3823-3828) provides a compelling picture of substrate-induced changes in the PcyA enzyme and the chemical basis of PcyA's catalytic activity. On the basis of these structures and the biochemical analysis of site-directed mutants of Nostoc PcyA, including mutants reported in recent studies (Tu et al. (2006) J. Biol. Chem. 281, 3127-3136) as well as mutants described in this study, a revised mechanism for the PcyA-mediated four-electron reduction of biliverdin IXalpha to 3E/3Z-phycocyanobilin via enzyme-bound bilin radical intermediates is proposed. The mechanistic insight of these studies, along with homology modeling, have provided new insight into the catalytic mechanisms of other members of the ferredoxin-dependent bilin reductase family that are widespread in oxygenic photosynthetic organisms.
Tu
Shih-Long
SL
Institute of Plant and Microbial Biology, Academia Sinica, Nankang, Taipei 115, Taiwan.
Rockwell
Nathan C
NC
Lagarias
J Clark
JC
Fisher
Andrew J
AJ
D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D008956:Models, Chemical; D016297:Mutagenesis, Site-Directed; D010088:Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017510:Protein Folding
D010088:Oxidoreductases; C071703:phytochromobilin synthase; C518487:HY2 protein, Arabidopsis; C518488:phycocyanobilin-ferredoxin oxidoreductase, Nostoc