Electrochemical genotoxicity screening for arylamines bioactivated by N-acetyltransferase.
Genotoxicity screening sensors that measure DNA damage from metabolism of arylamines were developed and evaluated. The sensors feature ultrathin films containing DNA and N-acetyltransferase (NAT) on pyrolytic graphite (PG) electrodes. NAT in the film catalyzed the conversion of the arylamine 2-aminofluorene (2-AF) to 2-acetylaminofluorene (2-AAF) by acetyl coenzyme A (AcCoA) dependent N-acetylation, as verified by liquid chromatography. DNA damage in the films from exposure to reactive 2-AF metabolites was measured subsequent to the enzyme reaction using catalytic voltammetric oxidation with Ru(bpy)32+. Square wave voltammetric (SWV) peaks increased with enzyme reaction time, and relative DNA damage rates at pH 5.8 were measured within 2 min. Control incubations of DNA/NAT films without AcCoA gave no significant sensor response. CapLC-MS/MS analysis of 2-AAF/DNA reaction products was consistent with 2-AF-guanine adducts formed in the films. DNA damage occurred more rapidly under weakly acidic conditions (pH 5.5-5.8) than at neutral pH, suggesting that genotoxicity from arylamine metabolism by NAT could be more significant in slightly acidic environments.
So
Minjeong
M
Department of Chemistry, 55 North Eagleville Road, University of Connecticut, Storrs, CT 06269, USA.
Hvastkovs
Eli G
EG
Bajrami
Besnik
B
Schenkman
John B
JB
Rusling
James F
JF
D015073:2-Acetylaminofluorene / Q000633:toxicity*; D000105:Acetyl Coenzyme A / Q000378:metabolism*; D000107:Acetylation; D001191:Arylamine N-Acetyltransferase / Q000378:metabolism*; D015374:Biosensing Techniques / Q000379:methods*; D002384:Catalysis; D002853:Chromatography, Liquid; D004247:DNA / Q000187:drug effects*; D004249:DNA Damage; D004563:Electrochemistry; D004566:Electrodes; D006108:Graphite / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D013058:Mass Spectrometry; D009152:Mutagenicity Tests / Q000379:methods*
D000105:Acetyl Coenzyme A; D006108:Graphite; D004247:DNA; D015073:2-Acetylaminofluorene; D001191:Arylamine N-Acetyltransferase