Purification of mixed-function amine oxidase from rat liver microsomes.
To clarify the metabolism of carcinogenic aminoazo dyes in target tissues, mixed function amine oxidase (MFAO) was purified from rat liver. The MFAO was solubilized from microsomes with Triton X in the presence of 20 glycerol and 1 mM EDTA and purified successively with DEAE Sepharose CL-6B, 2',5'-ADP Sepharose 4B and Hydroxyapatite column chromatography. The purified enzyme yielded a single protein band on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. The apparent molecular weight was about 59,000. When dimethylaniline (DMA) was used as a substrate, the specific activity of the enzyme fortified with NADPH was about 430 nmol DMA N-oxide formed/mg protein/min with a yield of about 15%. N-Demethylation of dimethylaminoazobenzene (DAB) with the enzyme proceeded only when iron was added to the reaction system.
D000818:Animals; D002845:Chromatography / Q000379:methods; D004492:Edetic Acid; D005990:Glycerol; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000302:isolation & purification* / Q000378:metabolism; D011092:Polyethylene Glycols; D051381:Rats; D011919:Rats, Inbred Strains; D004124:p-Dimethylaminoazobenzene / Q000378:metabolism
D011092:Polyethylene Glycols; D004492:Edetic Acid; D004124:p-Dimethylaminoazobenzene; D000587:Oxidoreductases Acting on CH-NH Group Donors; C019668:amine oxidase, mixed function; D005990:Glycerol