The effect of the length and flexibility of the side chain of basic amino acids on the binding of antimicrobial peptides to zwitterionic and anionic membrane model systems

The intent of this investigation was to determine the effect of varying the side chain length of the basic amino acids residues on the binding of a series of antimicrobial peptides (AMPs) to zwitterionic and anionic LUVs, SUVs and micelles. These AMPs are based on the incorporation of three dipeptide units consisting of the unnatural amino acids Tic-Oic in the sequence, Ac-GF-Tic-Oic-GX-Tic-Oic-GF-Tic-Oic-GX-Tic-XXXX-CONH₂, where X (Spacer #2) may be one of the following amino acids, Lys, Orn, Dab, Dpr or Arg. A secondary focus of this study was to attempt to correlate the possible mechanisms of membrane binding of these AMPs to their bacterial strain potency and selectivity. These AMPs produced different CD spectra in the presence of zwitterionic DPC and anionic SDS micelles. This observation indicates that these AMPs adopt different conformations on binding to the surface of zwitterionic and anionic membrane model systems. The CD spectra of these AMPs in the presence of zwitterionic POPC and anionic 4:1 POPC/POPG LUVs and SUVs also were different, indicating that they adopt different conformations on interaction with the zwitterionic and anionic liposomes. This observation was supported by ITC and calcein leakage data that indicated that these AMPs interact via very different mechanisms with anionic and zwitterionic LUVs. The enthalpy for the binding of these AMPs to POPC directly correlates to the length of Spacer #2. The enthalpy of binding of these AMPs to 4:1 POPC/POPG, however do not correlate with the length of Spacer #2. Clear evidence exists that the AMP containing the Dpr residues (the shortest length spacer) interacts very differently with both POPC and 4:1 POPC/POPG LUVs compared to the other four compounds. Data indicates that both the hydrophobicity and the charge distribution of Spacer #2, contribute to defining antibacterial activity. These observations have major implications on the development of these analogs as potential therapeutic agents.