FIG 3.

Effect of salt on SLA₈₀-NS5 binding. (A) Fluorescence titrations of fluorescein-labeled SLA₈₀ with NS5 (λ_ex = 480 nm; λ_em = 520 nm) in buffer B1 (50 mM Tris [pH 7.5] at 10°C, 1 mM MgCl₂) containing the following different NaCl concentrations: 80 mM, 100 mM, 130 mM, 170 mM, and 200 mM. The concentration of fluorescein-labeled SLA₈₀ is 1.5 × 10⁻⁸ M. Increasing salt concentrations decrease the affinity of the polymerase for SLA₈₀. The solid lines are nonlinear least-squares fits of the titration curve with a K of 7.0 × 10⁶ M⁻¹ (80 mM NaCl), a K of 5.5 × 10⁶ M⁻¹ (100 mM NaCl), a K of 4.0 × 10⁶ M⁻¹ (130 mM NaCl), a K of 3.5 × 10⁶ M⁻¹ (170 mM NaCl), and a K of 2.0 × 10⁶ M⁻¹ (200 mM NaCl). (B) Dependence of the logarithm of the intrinsic binding constant, K, on the logarithm of NaCl concentrations. The solid line is the linear least-squares fit of the NaCl concentration regions of the plot, which provide the slope, ∂logK/∂log[NaCl] = −1.3.