Partial purification and separation of multiple forms of cytochrome. P-450 and cytochrome P-448 from rat liver microsomes.
1. Partial purification of liver microsomal cytochrome p-450 results in the separation of two forms of cytochrome p-450 from phenobarbital-treated rats and two forms of cytochrome p-44, from 3-methylcholanthrene-treated rats. 2. Each of the four cytochrome fractions had different spectral properties (absolute spectra, CO differences spectra, and ethylisocyanide difference spectra). 3. The hemeprotein in fractions which elute from a DEAE-cellulose column at 100 mKM KCl fraction IV B) are more highly purified than the hemeproteins (fraction IV A) that elute in the column volume. 4. The more highly purified cytochrome fractions (IV B) contain 9-11 moles of cytochrome P-450 or P-448 per mg protein (an approximately 5-7 fold purification over microsomes) and are enzymatically active in the metabolism of a variety of substrates when combined with lipid and NADPH-cytochrome c reductase. These hemeprotein fractions are free of cytochrome b5 and NADPH-cytochrome c reductase, and the hemeproteins are purified approximately 100-fold with respect to phospholipid. The cytochrome P-450 and P-448 are virtually free of epoxide hydrase.
D000818:Animals; D001580:Benzopyrenes / Q000378:metabolism; D001589:Benzphetamine / Q000378:metabolism; D002248:Carbon Monoxide; D002848:Chromatography, DEAE-Cellulose; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis / Q000302:isolation & purification*; D003580:Cytochromes / Q000032:analysis / Q000302:isolation & purification*; D004851:Epoxide Hydrolases / Q000032:analysis; D004980:Ethane / Q000031:analogs & derivatives / Q000494:pharmacology; D008055:Lipids / Q000494:pharmacology; D008297:Male; D008748:Methylcholanthrene / Q000494:pharmacology; D008862:Microsomes, Liver / Q000201:enzymology* / Q000378:metabolism; D009097:Multienzyme Complexes; D009251:NADPH-Ferrihemoprotein Reductase / Q000032:analysis; D009281:Naphthalenes / Q000378:metabolism; D009570:Nitriles / Q000494:pharmacology; D010084:Oxidation-Reduction; D010634:Phenobarbital / Q000494:pharmacology; D010713:Phosphatidylcholines / Q000032:analysis; D051381:Rats; D013057:Spectrum Analysis; D013501:Surface-Active Agents / Q000494:pharmacology
D001580:Benzopyrenes; D003580:Cytochromes; D008055:Lipids; D009097:Multienzyme Complexes; D009281:Naphthalenes; D009570:Nitriles; D010713:Phosphatidylcholines; D013501:Surface-Active Agents; D001589:Benzphetamine; D008748:Methylcholanthrene; D002248:Carbon Monoxide; D003577:Cytochrome P-450 Enzyme System; D009251:NADPH-Ferrihemoprotein Reductase; D004851:Epoxide Hydrolases; D004980:Ethane; D010634:Phenobarbital