Lipid-binding and antimicrobial properties of synthetic peptides of bovine apolipoprotein A-II

We previously showed that bovine apolipoprotein A-II (apoA-II) had antimicrobial activity against Escherichia coli and the yeast Saccharomyces cere v isiae in PBS. We have characterized here the active domain of apoA-II using synthetic peptides. A peptide corresponding to C-terminal residues Leu 49 -Thr 76 exhibited significant antimicrobial activity against E. coli in PBS, but not against S. cere v isiae . Experiments using amino-acid-substituted peptides indicated that the residues Phe 52 -Phe 53 -Lys 54 -Lys 55 are required for the activity. Peptide Leu 49 -Thr 76 induced the release of calcein trapped inside the vesicles whose lipid composition resembles that of E. coli membrane, suggesting that peptide Leu 49 -Thr 76 can destabilize the E. coli membrane. CD measurements showed that the α-helicity of peptide Leu 49 -Thr 76 increased from 3.5 to 36% by addition of the vesicles. When E. coli cells were incubated with peptide Leu 49 -Thr 76 , some proteins were released to the external medium, probably owing to membrane destabilization caused by the peptide. In electron micrographs of E. coli cells treated with peptide Leu 49 -Thr 76 , transparent nucleoids and granulated cytoplasm were observed. Amino acid substitutions, Phe 52 Phe 53 → AlaAla (Phe 52,53 → Ala) in peptide Leu 49 -Thr 76 caused the loss of antimicrobial activity against E. coli, although protein-releasing activity was retained. Electron micrographs of the cells treated with peptide Leu 49 -Thr 76 (Phe 52,53 → Ala) revealed morphological change only at the nucleoids. Therefore peptide Leu 49 -Thr 76 appears to primarily target the cytoplasm rather than the membrane of E. coli cells.