FIGURE 2.

RP-HPLCs of sMamb-1 synthesis and refolding and MS spectrum and circular dichroism analyses. A, crude mixture obtained after SPPS of sMamb-1, and the major peak at 28.5 min corresponds to mambalgin-1; B, crude mixture after 36 h of refolding; and C, chromatogram of the purified toxin. Separations were performed on a X-Bridge BHE C18-300-5 analytical column (Waters) (250 × 4.5 mm; 1 ml·min⁻¹; solvent A, 0.1%H₂O/TFA; solvent B, 0.1%acetonitrile/TFA; gradient, 20–40% solvent B in 40 min). Inset in C shows the ESI-MS spectrum of the individual peak. D, circular dichroism spectra of wild-type and alanine variants of sMamb-1 showing their three-finger fold signature characterized by a maxima and a minima at 202 and 213 nm, respectively.