a) SDS-PAGE of the first ligation reaction between the C-peptide (C, residues 82–160) and the filter peptide (residues 70–81) to form the intermediate peptide (I, residues 70–160) at 0 min (lane 1) and 24 h (lane 2). b) ES-MS of the intermediate peptide. Inset, reconstructed spectrum (Expected mass = 11431.9). c) SDS-PAGE of the second ligation reaction between the N-peptide thioester (N, residues 1–69) and the intermediate peptide to form the KcsA polypeptide at 0 min (lane 1) and 24 h (lane 2). d) SDS-PAGE showing the folding of semi-synthetic KcsA by lipids. The unfolded monomeric (M, which corresponds to the KcsA polypeptide, P) and the folded tetrameric KcsA (T) are indicated.