ENHANCEMENT OF N-HYDROXY-2-AMINOFLUORENE BACTERIAL MUTAGENICITY BY THE SOLUBLE-PROTEIN FRACTION FROM RAT-LIVER AND PARTIAL-PURIFICATION OF THE ENHANCEMENT ACTIVITY

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  • 1 January 1981
    • journal article
    • research article
    • Vol. 41  (11) , 4600-4605
Abstract
Bacterial mutagenesis from 2-aminofluorene [AF, a carcinogen] mediated by washed rat liver microsomes was elevated 2- to 3-fold by addition of the hepatic soluble protein fraction. Enhancement was observed at 2 AF concentrations between 1-20 .mu.g/assay but not at 30-50 .mu.g/assay. The soluble protein fraction (without added microsomes) did not activate 2-aminofluorene for bacterial mutagenesis. Mutagenesis by N-hydroxy-2-aminofluorene or 2-nitrosofluorene was enhanced by the soluble protein fraction but only when NADPH was present. NADPH reduced 2-nitrosofluorene to N-hydroxy-2-aminofluorene and protected the hydroxylamine from oxidation, thus indicating that it was the mutagenicity of N-hydroxy-2-aminofluorene (but not 2-nitrosofluorene) which was enhanced by the soluble protein fraction. Without the added soluble protein fraction, mutagenesis by N-hydroxy-2-aminofluorene or 2-nitrosofluorene was unaffected by NADPH. A protein fraction with properties of the enhancement activity was partially purified. The fraction, which represents a 14-fold increase in specific activity, was assigned a MW of 33,500 by gel filtration through Sephadex G-100. This fraction was resolved into 3 components by polyacrylamide gel electrophoresis; the MW of the 3 components were determined by sodium dodecyl sulfate-polyacrylamide gel (10%) electrophoresis to be 33,000, 27,000 and 16,250. The mechanism of mutagenesis enhancement remains unknown.