Purification and biochemical characterization of reno-ferredoxin from bovine kidney mitochondria.
Bovine reno-ferredoxin was purified from kidney mitochondria by an improved method that included hydrophobic and ion-exchange chromatography on Toyopearl gels. The optical absorption spectrum of the oxidized reno-ferredoxin revealed two peaks, at 414 and 455 nm in the visible region. The minimum molecular weight of the ferredoxin was 12,900 Da by SDS-polyacrylamide gel electrophoresis. The amino acid residues of the NH2-terminal sequence of the ferredoxin were investigated using by a gas-phase sequencer. Bovine reno-ferredoxin and adreno-ferredoxin showed almost identical NH2-terminal amino acid sequences. Reconstitution of the 25-hydroxyvitamin D3-1 alpha -hydroxylase system was performed with the following three components: NADPH-ferredoxin reductase from bovine kidney mitochondria, reno-ferredoxin, and cytochrome P-450(D1 alpha) from bovine kidney mitochondria. The results demonstrated that the reno-ferredoxin was essential for the 1 alpha-hydroxylase activity of 25-hydroxyvitamin D3.
Yamazaki
M
M
Department of Legal Medicine, Osaka University, Medical School, Japan.
D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D005288:Ferredoxins / Q000737:chemistry / Q000302:isolation & purification*; D007668:Kidney / Q000737:chemistry* / Q000201:enzymology; D008928:Mitochondria / Q000737:chemistry / Q000201:enzymology; D008969:Molecular Sequence Data; D008970:Molecular Weight
D005288:Ferredoxins; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase