Figure 4.

In vitro characterization of EYFP–calmodulin insert protein. (A) Absorbance spectra with EDTA and zero Ca²⁺, or with saturating Ca²⁺, both at pH 7.5 and the same protein concentration. (B) Fluorescence excitation and emission spectra obtained under the same conditions. (C) Titration at pH 7.5 with Ca²⁺ buffers, plotting fractional conversion to the Ca²⁺-bound state vs. the logarithm of the free-Ca²⁺ concentration in molar units. (D) Titrations of normalized fluorescence vs. pH with EDTA and zero Ca²⁺, or with saturating Ca²⁺. The midpoints of the titration curves are 10.1 in EDTA and 8.9 in excess Ca²⁺.