Purification and characterization of rat pulmonary cytochrome P-450.
The pulmonary cytochrome P-450, P450 L-2, was purified 460-fold from pulmonary microsomes of untreated male rats. Its specific content was 10.6 nmol/mg of protein. The monomeric molecular weight was 54,000 on SDS-polyacrylamide gel electrophoresis. The CO-reduced absorption maximum of P450 L-2 was at 451 nm, and the oxidized heme iron appeared to be in the low-spin state, as deduced from the Soret maximum at 421 nm. P450 L-2 had high lauric acid omega- and (omega-1)-hydroxylation activities, but low prostaglandin A1 omega- and (omega-1)-hydroxylation activities. It catalyzed the O-dealkylation of 7-ethoxycoumarin, but was not efficient in the hydroxylation of testosterone or the N-demethylation of aminopyrine. The NH2-terminal amino acid sequence of P450 L-2 was V-L-N-F-L-X-P-X-L (X being an unidentified residue). The catalytic properties of P450 L-2 resembled those of P450 K-5, the major rat renal cytochrome P-450. However, anti-P450 K-5 antibody did not cross-react with P450 L-2, and these forms had different NH2-terminal sequences. To judge from the results of NH2-terminal sequence analysis, P450 L-2 seems to be placed in the IVB gene family. Also, P-450 IIB1 was detected by immunoblotting in one of the peaks on ion-exchange HPLC during the purification of P450 L-2, suggesting the presence of P-450 IIB1 in rat pulmonary microsomes.
Imaoka
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Laboratory of Chemistry, Osaka City University Medical School.
Funae
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Y
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002384:Catalysis; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008168:Lung / Q000201:enzymology*; D008297:Male; D008861:Microsomes / Q000201:enzymology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D051381:Rats; D011919:Rats, Inbred Strains
D003577:Cytochrome P-450 Enzyme System