Figure 4

Visible Circular dichroism (CD) spectra of Ni²⁺ bound to WT and histidine mutant proteins.

(A) Visible CD spectra (300–600 nm) of Ni²⁺ bound to 20 μM WT in 150 mM NaCl, 25 mM potassium phosphate,pH 7.4, 1 mM DTT and 0.3% glycerol with 50 and 250 molar equivalents of EDTA. The differences in ellipticity were approximated at 440 and 538 nm. (B) Visible CD spectra of Ni²⁺ bound to 20 μM WT and histidine mutants. The differences in ellipticity values were approximated at 440 and 538 nm, indicating different amounts of Ni²⁺ binding to the proteins. Data are representative of three independent experiments. (C) Intensities of the negative band at 538 nm of WT and histidine mutants. Data were normalized and compared to WT (whereby intensity of WT was defined as 100%).