Structure/function studies of an endotoxin-neutralizing peptide derived from bactericidal/permeability-increasing protein.
BACKGROUND
Bactericidal/permeability-increasing protein, BPI, has a beta-turn with alternating cationic and hydrophobic residues in its lipopolysaccharide (endotoxin, LPS)-binding domain. A peptide, betapep25, was designed with 9 residues of the LPS-binding domain of BPI flanked by beta-turn-inducing elements. Thereafter, we sought to use single amino acid substitutions to identify residues that are important for the biological activities of betapep25.


METHODS
Single alanine or norleucine replacement "walkthrough" peptides based on betapep25 were generated and tested for their ability to kill P aeruginosa and to neutralize endotoxin.


RESULTS
Substitution of all lysines inhibited bactericidal activity. Inhibition of LPS-neutralizing activity was seen in 9 peptides in which an alanine or norleucine was substituted for each of 4 of the basic residues and 1 hydrophobic residue from the LPS-binding region of BPI and 4 hydrophobic residues from the beta-turn-inducing regions flanking the LPS-binding region on the carboxy-terminal side. Intriguingly, these last 4 substitutions resulted in peptides that exhibited increased bactericidal activity compared to betapep25.


CONCLUSIONS
These results demonstrate the importance of both cationic and hydrophobic amino acid residues to bactericidal and endotoxin-neutralizing activities. These perturbations of biological activity should be considered in the design of synthetic peptide endotoxin antagonists.
Wasiluk
Karen R
KR
Departments of Surgery and Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, Minn USA.
Leslie
Daniel B
DB
Vietzen
Paul S
PS
Mayo
Kevin H
KH
Dunn
David L
DL
D000595:Amino Acid Sequence; D000818:Animals; D023181:Antimicrobial Cationic Peptides; D001419:Bacteria / Q000187:drug effects; D001798:Blood Proteins / Q000494:pharmacology*; D002460:Cell Line; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors*; D008565:Membrane Proteins / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D013329:Structure-Activity Relationship; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000096:biosynthesis
D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D008070:Lipopolysaccharides; D008565:Membrane Proteins; D014409:Tumor Necrosis Factor-alpha; C021623:bactericidal permeability increasing protein