Identification and heterologous expression of the cytochrome P450 oxidoreductase from the white-rot basidiomycete Coriolus versicolor.
A cDNA encoding cytochrome P450 oxidoreductase (CPR) from the lignin-degrading basidiomycete Coriolus versicolor was identified using RT-PCR. The full-length cDNA consisted of 2,484 nucleotides with a poly(A) tail, and contained an open reading frame. The G+C content of the cDNA isolated was 60%. A deduced protein contained 730 amino acid residues with a calculated molecular weight of 80.7 kDa. The conserved amino acid residues involved in functional domains such as FAD-, FMN-, and NADPH-binding domains, were all found in the deduced protein. A phylogenetic analysis demonstrated that C. versicolor CPR is significantly similar to CPR of the basidiomycete Phanerochaete chrysosporium and that they share the same major branch in the fungal cluster. A recombinant CPR protein was expressed using a pET/ Escherichia coli system. The recombinant CPR protein migrated at 81 kDa on SDS polyacrylamide gel electrophoresis. It exhibited an NADPH-dependent cytochrome c reducing activity.
Ichinose
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Faculty of Agriculture, Kyushu University, Fukuoka, Japan.
Wariishi
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Tanaka
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D000595:Amino Acid Sequence; D001483:Base Sequence; D001487:Basidiomycota / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D003574:Cytochrome c Group / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009251:NADPH-Ferrihemoprotein Reductase / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D010802:Phylogeny; D012331:RNA, Fungal / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D003574:Cytochrome c Group; D018076:DNA, Complementary; D012331:RNA, Fungal; D011994:Recombinant Proteins; D009251:NADPH-Ferrihemoprotein Reductase