Figure 4.

Titration of Rox–40-1, SgrAI, with Flo–PC DNA in the presence of 10 mm Ca²⁺. A, 50 nm Rox–40-1, 2 μm SgrAI, 10 mm CaCl₂, and buffer A (see “Experimental procedures”) with PC (1:9 Flo–PC:PC) at 25 °C. Emission spectra taken with 498-nm excitation, and corrected for Flo emission, dilution, and Rox emission caused by absorption at 498 nm. Concentration of total EP₄₀ (SgrAI bound to two Flo–PC molecules, calculated as half the concentration of total added Flo–PC and assumes complete binding of Flo–PC) shown in nm. B, average corrected intensities from A (wavelengths 603–613 nm) versus total EP₄₀ (the concentration of SgrAI bound to Flo–PC DNA) (filled blue circles). Values are fit to the Hill equation (red line, r = 0.9985) to give K_½ = 0.16 ± 03 μm and Hill coefficient = 1.1 ± 0.1.