Monoclonal antibodies to liver microsomal cytochrome P-450E of the marine fish Stenotomus chrysops (scup): cross reactivity with 3-methylcholanthrene induced rat cytochrome P-450.
Hybridomas were prepared from myeloma cells and spleen cells of BALB/c female mice immunized with hepatic cytochrome P-450E purified from the marine fish, Stenotomus chrysops (scup). Nine independent hybrid clones produced MAbs, either IgG1, IgG2b, or IgM, that bound to purified cytochrome P-450E in radioimmunoassay. Antibodies from one clone MAb (1-12-3), also strongly recognized rat cytochrome P-450MC-B (P-450BNF-B; P-450c). The nine antibodies inhibited reconstituted aryl hydrocarbon hydroxylase (AHH) and ethoxycoumarin O-deethylase of scup cytochrome P-450E to varying degrees, and inhibited AHH activity of beta-naphthoflavone-induced scup liver microsomes in a pattern similar to that in reconstitutions, indicating that cytochrome P-450E is identical to the AHH catalyst induced in this fish by beta-naphthoflavone. MAb 1-12-3 also inhibited the reconstituted AHH activity of the major BNF-induced rat isozyme. Conversely, MAb 1-7-1 to rat cytochrome P-450MC-B had little effect on AHH activity of scup cytochrome P-450E, and did not recognize cytochrome P-450E in radioimmunoassay nor in an immunoblot. Scup cytochrome P-450E and rat cytochrome P-450MC-B thus have at least one common epitope recognized by MAb 1-12-3, but the epitope recognized by Mab 1-7-1 is absent or recognized with low affinity in cytochrome P-450E. The various assays indicate that the nine MAbs against cytochrome P-450E are directed to different epitopes of the molecule. These MAbs should be useful in determining phylogenetic relationships of the BNF- or MC-inducible isozymes and their regulation by other environmental factors.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D003429:Cross Reactions; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology*; D004790:Enzyme Induction / Q000187:drug effects; D000939:Epitopes / Q000032:analysis; D005399:Fishes / Q000378:metabolism*; D005779:Immunodiffusion; D007527:Isoenzymes / Q000276:immunology*; D008748:Methylcholanthrene / Q000494:pharmacology; D051379:Mice; D008862:Microsomes, Liver / Q000201:enzymology*; D011817:Rabbits; D011863:Radioimmunoassay; D051381:Rats
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007527:Isoenzymes; D008748:Methylcholanthrene; D003577:Cytochrome P-450 Enzyme System