Chemical Synthesis and Characterization of an Equinatoxin II(1–85) Analogue

Abstract

The chemical synthesis of an 85 residue analogue of the pore-forming protein, Equinatoxin II (EqtII), was achieved. Peptide precursors with over 40 residues were assembled by solid phase synthesis. The EqtII(1–46) fragment was modified to the reactive C-terminal thioester and native chemical ligation was performed with the A47C mutated EqtII(47–85) peptide to form the EqtII(1–85) analogue. Circular dichroism spectroscopy indicated that the N-terminal domain of EqtII(1–46) and EqtII(1–85) maintains predominantly an α-helical structure in solution and also in the presence of lipid micelles. This demonstrates the feasibility of assembling the full 179 residue protein EqtII via chemical means. Site-specific isotopic labels could be incorporated for structural studies in membranes by solid-state NMR spectroscopy.

Keywords

• native chemical ligation
• solid-phase peptide synthesis
• selectively labelled proteins
• membrane protein structure
• solid-state NMR