Uridine diphosphate-N-acetylglucosamine (UNAG)-dependent ζ hydrolysis of ATP is poorly competed by GTP. Samples containing 2 mM UNAG and increasing concentrations of ATP (0.5, 5 and 10 mM (with a fixed concentration of α32P-ATP, 10 nM), lanes 2–4) or 2 mM UNAG and a fixed concentration of ATP (0.5 mM [α32P-ATP, 10 nM]) and increasing GTP concentrations (1.25, 2.5, 5, 7.5 and 10 mM, lanes 5–9) were incubated with 0.5 µM ζ toxin for 30 min at 30 °C in buffer B. ATP hydrolysis was analyzed by thin-layer chromatography (TLC) performed on polyethyleneimine-cellulose plates with 0.85 M KH2PO4 (pH 3.4) as the mobile phase, followed by autoradiography.