Binary and Ternary Binding Affinities between Exonuclease-Deficient Klenow Fragment (Kf-exo<sup>–</sup>) and Various Arylamine DNA Lesions Characterized by Surface Plasmon Resonance

We used surface plasmon resonance (SPR) to characterize the binding interactions between the exonulease-free Klenow fragment (Kf-exo<sup>–</sup>) and unmodified and modified dG adducts derived from arylamine carcinogens: fluorinated 2-aminofluorene (FAF), 2-acetylaminofluorene (FAAF), and 4-aminobiphenyl (FABP). Tight polymerase binding was detected with unmodified dG and the correct dCTP. The discrimination of correct versus incorrect nucleotides was pronounced with <i>K</i><sub>D</sub> values in the order of dCTP ≪ dTTP < dATP < dGTP. In contrast, minimal selectivity was observed for the modified templates with Kf-exo<sup>–</sup> binding tighter to the FAAF (<i>k</i><sub>off</sub>: 0.02 s<sup>–1</sup>) and FABP (<i>k</i><sub>off</sub>: 0.01 s<sup>–1</sup>) lesions than to FAF (<i>k</i><sub>off</sub>: 0.04 s<sup>–1</sup>).