Sequence motif-specific assignment of two [2Fe-2S] clusters in rat xanthine oxidoreductase studied by site-directed mutagenesis.
The sequence motif-specific assignment of the two distinct [2Fe-2S] clusters in rat xanthine oxidoreductase (XOR) was unequivocally established by site-directed mutagenesis of recombinant enzymes expressed in a baculovirus-insect cell system and electron paramagnetic resonance (EPR) spectroscopy. The conserved cysteine residues, including Cys-115, in the unusual C-terminal -Cys-Xaa(2)-Cys-//-Cys-Xaa(1)-Cys- motif serve as ligands to the Fe/S I center, which is probably located in close proximity to the Mo-pterin center. Other conserved cysteine residues, including Cys-43 and Cys-51, in the N-terminal plant ferredoxin-like motif serve as ligands to the Fe/S II center, which is distantly located from the Mo-pterin center. The present sequence motif-specific assignment of the Fe/S I and II centers is discussed in the light of the structural features of XOR.
Iwasaki
T
T
Department of Biochemistry and Molecular Biology, Nippon Medical School, Sendagi, Tokyo 113-8602, Japan.
Okamoto
K
K
Nishino
T
T
Mizushima
J
J
Hori
H
H
D000595:Amino Acid Sequence; D000818:Animals; D017124:Conserved Sequence; D003545:Cysteine; D004578:Electron Spin Resonance Spectroscopy; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D051381:Rats; D011994:Recombinant Proteins; D014968:Xanthine Dehydrogenase / Q000737:chemistry* / Q000235:genetics; D014969:Xanthine Oxidase / Q000737:chemistry* / Q000235:genetics
D007506:Iron-Sulfur Proteins; D011994:Recombinant Proteins; D014968:Xanthine Dehydrogenase; D014969:Xanthine Oxidase; D003545:Cysteine