Immunochemical and sub-structural characterization of sheep lung cytochrome P450LgM2.
Sheep lung cytochrome P450LgM2 belonging to gene subfamily 2B, was obtained in highly purified form and antibodies against sheep lung cytochrome P450LgM2 were produced in rabbits by using the previously developed methods in our laboratory. Immunological and enzymatic studies showed that antibodies against lung cytochrome P450LgM2 inhibited benzphetamine N-demethylation, ethylmorphine N-demethylation and aniline 4-hydroxylation reactions in sheep lung microsomes about 99, 80 and 62%, respectively. Benzphetamine N-demethylation reaction in sheep lung microsomes was only catalyzed by cytochrome P450LgM2 isozyme while the other isozymes of P450 as well as P450LgM2 are also involved in the metabolism of ethylmorphine and aniline. Similar to lung microsomes, benzphetamine N-demethylase activity of the reconstituted systems containing purified sheep lung cytochrome P450LgM2 or phenobarbital (PB)-treated rabbit liver cytochrome P450LM2(2B4) was also inhibited by P450LgM2 antibodies about 95 and 82%, respectively. A 50% inhibitory effect of sheep P450LgM2 antibodies was also observed in ethylmorphine N-demethylase activity of the reconstituted system containing purified sheep lung cytochrome P450LgM2. SDS-PAGE peptide maps obtained following the partial proteolysis of purified sheep lung cytochrome P450LgM2 and PB-rabbit liver P450LM2(2B4) isozymes, using chymotrypsin and papain, were similar in general. However they showed some differences both qualitatively and quantitatively, suggesting that some differences exist among the amino acid sequences of sheep lung cytochrome P450LgM2 and rabbit liver cytochrome P4502B4.
Adali
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O
Department of Biology, Middle East Technical University, Ankara, Turkey.
Abu-Baker
T
T
Arinç
E
E
D000818:Animals; D000918:Antibody Specificity; D015153:Blotting, Western; D003577:Cytochrome P-450 Enzyme System; D007120:Immunochemistry; D008168:Lung / Q000201:enzymology* / Q000648:ultrastructure; D008861:Microsomes / Q000201:enzymology*; D010089:Oxidoreductases, N-Demethylating / Q000737:chemistry* / Q000378:metabolism; D010449:Peptide Mapping; D011817:Rabbits; D012756:Sheep; D013045:Species Specificity
D003577:Cytochrome P-450 Enzyme System; C019781:benzphetamine N-demethylase; D010089:Oxidoreductases, N-Demethylating