Effects of Single Amino Acid Substitution on the Biophysical Properties and Biological Activities of an Amphipathic α-Helical Antibacterial Peptide Against Gram-Negative Bacteria

An antimicrobial peptide, known as V13K, was utilized as the framework to study the effects of charge, hydrophobicity and helicity on the biophysical properties and biological activities of alpha-helical peptides. Six amino acids (Lys, Glu, Gly, Ser, Ala, and Leu) were individually used to substitute the original hydrophobic valine at the selected sixteenth location on the non-polar face of V13K. The results showed that the single amino acid substitutions changed the hydrophobicity of peptide analogs as monitored by RP-HPLC, but did not cause significant changes on peptide secondary structures both in a benign buffer and in a hydrophobic environment. The biological activities of the analogs exhibited a hydrophobicity-dependent behavior. The mechanism of peptide interaction with the outer membrane and cytoplasmic membrane of Gram-negative bacteria was investigated. We demonstrated that this single amino acid substitution method has valuable potential for the rational design of antimicrobial peptides with enhanced activities.