Secondary structure and stability of CaHD. A) Full CD spectra of 0.05 mg/mL of CaHD were collected every 10°C during a temperature melt with a temperature ramp of 1 °C/min. B) The changes in the mean residue ellipticity (MRE) of CaHD at 222 nm as a function of temperature. C) Differential scanning calorimetry of 1 mg/mL purified CaHD (50 mM sodium phosphate, pH 7.4, 200 mM NaCl) using an increasing temperature ramp of 0.5 °C/min. D) A single low-energy transition at 57 °C is observed. Data were fit to a two-state transition.