Sarcotoxin IA is a 39-residue cecropin-type peptide from Sarcophaga peregrina. This peptide exhibits antibacterial activity against Gram-negative bacteria through its interaction with lipid A, a core component of lipopolysaccharides. To acquire detailed structural information on this specific interaction, we performed NMR analysis using bacterially expressed sarcotoxin IA analogs with C-13- and N-15-labeling along with lipid A-embedding micelles composed of dodecylphosphocholine. By inspecting the stable isotope-assisted NMR data, we revealed that the N-terminal segment (Leu3-Arg18) of sarcotoxin IA formed an amphiphilic alpha-helix upon its interaction with the aqueous micelles. Furthermore, chemical shift perturbation data indicated that the amino acid residues displayed on this alpha-helix were involved in the specific interaction with lipid A. On the basis of these data, we successfully identified Lys4 and Lys5 as key residues in the interaction with lipid A and the consequent antibacterial activity. Therefore, these results provide unique information for designing chemotherapeutics based on antibacterial peptide structures. (C) 2013 Elsevier Inc. All rights reserved.