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

• Published in: Biochemical journal 1999-08, Vol.342 ( Pt 1) (1), p.215-221
• Main Authors: Motizuki, M, Itoh, T, Satoh, T, Yokota, S, Yamada, M, Shimamura, S, Samejima, T, Tsurugi, K
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amino Acid Substitution; Animals; Anti-Bacterial Agents; Anti-Infective Agents - chemistry; Anti-Infective Agents - isolation & purification; Anti-Infective Agents - metabolism; Anti-Infective Agents - pharmacology; Apolipoprotein A-II - chemistry; Apolipoprotein A-II - isolation & purification; Apolipoprotein A-II - metabolism; Apolipoprotein A-II - pharmacology; Bacterial Proteins - chemistry; Bacterial Proteins - metabolism; Cattle; Cell Membrane - drug effects; Cell Membrane - metabolism; Circular Dichroism; Cytoplasm - drug effects; Cytoplasm - metabolism; Escherichia coli - cytology; Escherichia coli - drug effects; Escherichia coli - metabolism; Escherichia coli - ultrastructure; Fluoresceins - metabolism; Inhibitory Concentration 50; Microscopy, Electron; Molecular Sequence Data; Molecular Weight; Peptide Fragments - chemical synthesis; Peptide Fragments - chemistry; Peptide Fragments - metabolism; Peptide Fragments - pharmacology; Phospholipids - metabolism; Protein Binding; Protein Structure, Secondary; Saccharomyces cerevisiae - cytology; Saccharomyces cerevisiae - drug effects; Sodium Chloride - pharmacology
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/0264-6021:3420215

We previously showed that bovine apolipoprotein A-II (apoA-II) had antimicrobial activity against Escherichia coli and the yeast Saccharomyces cerevisiae 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. cerevisiae. 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 alpha-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.