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 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.
Motizuki
M
M
Department of Biochemistry 2, Yamanashi Medical University, Yamanashi 409-3898, Japan.
Itoh
T
T
Satoh
T
T
Yokota
S
S
Yamada
M
M
Shimamura
S
S
Samejima
T
T
Tsurugi
K
K
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology*; D016633:Apolipoprotein A-II / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002942:Circular Dichroism; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D004926:Escherichia coli / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D005452:Fluoresceins / Q000378:metabolism; D020128:Inhibitory Concentration 50; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D010743:Phospholipids / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000187:drug effects; D012965:Sodium Chloride / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D000890:Anti-Infective Agents; D016633:Apolipoprotein A-II; D001426:Bacterial Proteins; D005452:Fluoresceins; D010446:Peptide Fragments; D010743:Phospholipids; D012965:Sodium Chloride; C007740:fluorexon