Specific labelling of microsomal proteins by reactive intermediates generated from 2-acetylaminofluorene in vitro |
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Authors: | Mustak Ali Kaderbhai Tony K. Bradshaw Robert B. Freedman |
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Affiliation: | 1. Biological Laboratory, University of Kent, Canterbury, CT2 7NJ Kent United Kingdom;2. Shell Toxicology Laboratory, Sittingbourne Research Centre, Sittingbourne, ME9 8AG United Kingdom |
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Abstract: | Incubation of 2-[9-14C] acetylaminofluorene (2-[9-14C]AAF) in vitro with rat liver microsomes, leads to covalent binding of label to microsomal proteins. The binding is NADPH-dependent, increases linearly with time, and is inhibited by SKF-525A and 7,8-benzoflavone (7,8-BF). Binding is increased more than 8-fold in microsomes from 3-methylcholanthrene(MC)-pretreated rats, but only less than 2-fold in those from phenobarbital(PB)-pretreated rats. In the presence of cytosolic proteins, there is slight enhancement of the labelling of microsomes and some labelling of the cytosolic proteins. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and 2-dimensional gel electrophoresis indicate that covalent labelling by 2-AAF derivatives is concentrated in specific proteins. The pattern of labelling varies between microsomes from animals pretreated with PB, MC and 2-AAF. Factors which may contribute to the specificity of labelling are discussed. |
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Keywords: | 2-AAF 2-acetylaminofluorene 7,8-BF 7,8-benzoflavone dichlorovos ETCP 1,2 epoxy 3,3,3-trichloropropane MC 3-methylcholanthrene PB phenobarbital SDS-PAGE sodium dodecyl sulphate polyacrylamide gel electrophoresis TCA trichloroacetic acid |
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