Purification and NH2-terminal amino acid sequences of human and rat kidney fatty acid omega-hydroxylases.
A cytochrome P-450 (P-450), designated P-450HK omega, has been isolated and purified from human kidney microsomes to a specific content of 13 nmoles of P-450/mg of protein. P-450HK omega showed an apparent molecular weight of 52,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Absolute spectra of the oxidized form indicated that this P-450 was largely in the low-spin state and partly in the high-spin state. It catalyzed the omega- and (omega-1)-hydroxylation of fatty acids such as laurate, myristate, and palmitate, with no activity toward prostaglandin A1, benzphetamine, 7-ethoxycoumarin, or 7-ethoxyresorufin. The first 35 NH2-terminal amino acid sequence of P-450HK omega had about 70% homology with those of rabbit kidney fatty acid omega-hydroxylases of the P-450 IVA gene subfamily, P-450ka-1, P-450ka-2, and P-450kd, except for four undetermined residues. Moreover, Western blot and immuno-inhibition studies showed that P-450HK omega reacted with an antibody against the rabbit kidney fatty acid omega-hydroxylase. The results suggest that P-450HK omega is a member of the same P-450 gene family (IVA subfamily) as the rabbit enzymes. In addition, the terminal sequence of P-450HK omega also showed 54% homology with that of P-450k-2, a fatty acid omega-hydroxylase from rat kidney microsomes. To our knowledge, this is the first time that a P-450 specific for fatty acid omega-hydroxylase activity has been isolated to homogeneity from human tissues.
Kawashima
H
H
Department of Urology, Osaka City University Medical School, Japan.
Kusunose
E
E
Kubota
I
I
Maekawa
M
M
Kusunose
M
M
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000302:isolation & purification*; D005227:Fatty Acids / Q000378:metabolism*; D005260:Female; D006801:Humans; D006900:Hydroxylation; D007668:Kidney / Q000201:enzymology*; D008297:Male; D008861:Microsomes / Q000737:chemistry / Q000201:enzymology; D008875:Middle Aged; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000302:isolation & purification*; D051381:Rats; D012689:Sequence Homology, Nucleic Acid
D005227:Fatty Acids; D010446:Peptide Fragments; D003577:Cytochrome P-450 Enzyme System