‘Lollipop’-shaped helical structure of a hybrid antimicrobial peptide of temporin B-lipopolysaccharide binding motif and mapping cationic residues in antibacterial activity

Abstract

BackgroundTemporins are attractive templates for the development of antibiotics. However, many temporins are inactive against Gram-negative bacteria. Previously, we demonstrated conjugation of a lipopolysaccharide binding motif peptide to temporins yielded hybrid non-haemolytic AMPs that killed several Gram-negative bacteria. MethodsWe carried out a systematic Ala replacement of individual cationic and polar amino acid residues of LG21, a hybrid AMP consisted of temporin B (TB) and LPS binding motif. These Ala containing analogs of LG21 were examined for antibacterial activity, cell membrane permeabilization and liposome leakage assays using optical spectroscopic methods. Atomic resolution structure of LG21 was determined in zwitterionic dodecyl phosphocholine (DPC) micelles by NMR spectroscopy. ResultsCationic residues in the LPS binding motif of LG21 were critical for bactericidal and membrane permeabilization. Detergent bound structure of LG21 revealed helical conformation containing extensive sidechain/sidechain packing including cation/π interactions in the LPS binding motif. The helical structure of LG21 resembled a ‘lollipop’ like shape that was sustained by a compacted bulky aromatic/cationic head with a comparatively thinner ‘stick’ at the N-terminal region. The ‘head’ of the structure could be localized into micelle-water interfacial region whereas the ‘stick’ region may be inserted into the hydrophobic core of micelle. ConclusionsThe LPS binding motif of LG21 played dominant roles in broad spectrum activity and the 3-D structure provided plausible mechanistic insights for permeabilization of bacterial membrane. General significanceHybrid AMPs containing LPS binding motif could be useful for the structure based development of broad spectrum antibiotics.