Figure 1 - Unfolding transition of trypsinogen in 100 mM MES, pH 7.0, 20 mM CaCl2 and 81 mM NaNO3, pH 7.0, as a function of GdnHCl concentration. The fraction of unfolded protein (fU) was calculated from the ratio of differential spectroscopy (DS) signal at 293 nm (triangles) and molar ellipticity ([Q]) at 300 nm (black circles) and at 222 nm (open circles). The fit for a two-state transition is represented by the dashed line for DS 293 nm and CD 300 nm and by the dotted line for CD 222 nm. The fits were calculated according to Santoro and Bolen (24). The three-state fit of CD at 222 nm (solid line) was calculated according to Matthews and Crisanti (25). The near-UV CD was monitored at 300 nm using a 0.5-cm path length cuvette. The spectroscopic measurements were made in a 0.5-cm path length tandem cell as described in the procedures. Protein concentration was 1 mM for each sample. The far-UV CD was monitored at 222 nm using a 0.1-cm path length cuvette. Protein concentration was 250 ÁM and temperature 25oC.