Figure 6.

Equilibrium urea unfolding of W79F M-TTR. (A) Fluorescence spectra at 3 µM protein concentration and urea concentration ranging from 0 to 7.8 M in 20 mM phosphate buffer at pH 7.4, 25 °C (excitation 290 nm, slits 5 and 10 nm for ex and em, respectively). (B) Urea denaturation curve using W79F M-TTR fluorescence at 330 nm. The solid line through the data represents the best fit to a two-state model (Santoro and Bolen, 1988). The obtained $\mathrm{\Delta }{G}_{U-F}^{H20}$ value is shown. (C) Corresponding second derivative spectra with arrows indicating the three major peaks. (D) Values of second derivative at 324, 346 and 367 nm versus urea concentration, plotted to obtain urea denaturation curves. The solid lines through the data represent the best fit to a two-state model (Santoro and Bolen, 1988). The obtained $\mathrm{\Delta }{G}_{U-F}^{H20}$ values are shown.