N-Acetyltransferase activities with p-aminobenzoic acid and 2-aminofluorene as substrates were determined in isolates of the bacterium Escherichia coli. The N-acetyltransferase activity was determined by an acetyl CoA recycling assay and high pressure liquid chromatography. The N-acetyltransferase activities from a number of E. coli isolates were found to be 0.67 +/- 0.04 nmole/min/mg protein for 2-aminofluorene, and 0.46 +/- 0.02 nmole/min/mg protein for p-aminobenzoic acid. The apparent K-m and V-max values obtained were 2.85 +/- 0.65 mM and 7.51 +/- 0.86 nmol/min/mg protein, respectively, for 2-aminofluorene, and 2.35 +/- 0.39 mM and 9.43 +/- 0.78 nmol/min/mg protein, respectively, for p-aminobenzoic acid. The optimal pH value for the enzyme activity was 7.0 for both substrates tested. The optimal temperature for enzyme activity was 37 degrees C for both substrates. The N-acetyltransferase activity was inhibited by iodoacetamide: at 0.25 mM iodoacetamide, activity was reduced 50%, and at 1.0 mM, more than 90%. Among a series of divalent cations and salts, CU2+ and Zn2+ were demonstrated to be the most potent inhibitors. This report is the first demonstration of acetyl CoA:arylamine N-acetyltransferase activity in E. coli.