Research

Aromatic amines is an important group of mutagens that includes Nitrofluorene, 2-Aminofluorene, 4-Aminobiphenyl etc. They are known to form C-8 substituted dG-adducts. For example, AF [N-(2′-deoxyguanosin-8-yl)-2-aminofluorene] and AAF [N-(2′-deoxyguanosin-8-yl)-2-acetylaminofluorene] are the two major dG-adducts formed by 2-nitrofluorene and 2-aminofluorene. These adducts are known to induce discrete biological outcomes i.e. mutation and repair. The NarI sequence (5′-G1G2CG3CC-3′) is a mutational hot spot for frameshift mutagenesis, where arylamine adduction at G3 exerts a high frequency of -2 deletion mutations. We have shown that the thermodynamic stability of bulged-out slipped mutagenic intermediates (SMI) is directly related to deletion mutations. Acetylaminofluorene is a prototype arylamine carcinogen that forms C8-dG-AAF and C8-dG-AF as the major DNA lesions. There have been conflicting NMR results as to which flanking cytosine (5ꞌ or 3ꞌ) loops out with modified-dG to form a SMI during translesion synthesis: (i) 3′-flanking dC loops out with AAF modified- dG31 (ii) 5′- flanking dC loops out with AF modified-dG32. Here we investigated the AF and AAF induced -2 bulge structures using fluorescence spectroscopy by substituting the flanking dC with a fluorescent analog pyrrolo-deoxycytidine (PC). We prepared two NarI templates; 5′–G1G2PCG3CC–3′ and 5′-G1G2CG3PCC-3′ with PC located at 5′ or 3′ flanking position to G3, respectively. Our results suggest that AAF prefers 3′-PC whereas AF favors 5′- PC in forming -2 deletion bulge structures, consistent with the above-referenced NMR studies. Overall, these results indicate that the ‘N-acetyl’ factor (e.g., N-acetylated AAF vs. N-deacetylated AF) plays a key role in inducing different types of bulge structures.
Related People: Bongsup Cho
Dept. of Biomedical and Pharmaceutical Sciences, College of Pharmacy, University of Rhode Island.