CD spectra of apo- and holoNCS in MG-like and native state. CD spectra of the natural holoNCS (dotted line) in 10 mM phosphate buffer (pH 3) and the constituted holoNCS of NCS-Chr and MG-like state of apoNCS (full line) in 1.2 M GdnHCl (pH 3). MG-like state of apoNCS for constitution was prepared by incubating 15 μM apoNCS with 1.2 M GdnHCl in 10 mM phosphate (pH 3) at 25°C for 30 min. Constitution was performed prior to CD measurement by adding 1:1 molar ratio of methanol extracted NCS-Chr into aqueous solution containing MG-like state of apoNCS. Final methanol content was kept minimal to about 4% (v/v).Inset: Near-UV CD spectrum of apoNCS in MG-like state (full line) (in the presence of 1.2 M GdnHCl (pH 3)) and in folded state (dotted line) (in the absence of GdnHCl in 10 mM phosphate buffer (pH 3)). All CD spectra were recorded in mean residue ellipticity at 25°C.