Studies On The Structure-activity Relationship And Action Mechanism Of Antimicrobial Peptides

Antimicrobial peptides (also known as host defense peptides) are evolutionarilyconserved components of the innate immune response and are found among allclasses of life. These peptides present different mechanism of action againstprokaryotic and eukaryotic. They are potent, and broad spectrum antibiotics whichexhibit potential as novel therapeutic agents. Antimicrobial peptides have beendemonstrated to kill Gram-negative and Gram-positive bacteria (including resistantstrains against conventional antibiotics), mycobacteria (including Mycobacteriumtuberculosis), fungi and even transformed or cancerous cells. Unlike the majority ofconventional antibiotics, antimicrobial peptides may also have the ability to enhanceimmunity by functioning as immunomodulators. In this study, we focus on thestructure-function relationships of a series of antimicrobial peptides and explore theaction mechanism of antimicrobial peptide by the relationships of biophysical activityand biological activity of peptide analogs.First, the26-residue α-helix peptide V13K was utilized as the framework tostudy the effects of peptide charge, hydrophobicity and helicity on peptide biologicalactivities. Six amino acid residues (Glu, Lys, Ser, Gly, Ala, and Leu) wereindividually used to substitute original hydrophobic valine at the selected sixteenthposition on the nonpolar face of peptide P. In order to analyse the effect of singleamino acid substitution on the biophysical activity and biological activity of thepeptides, the selected position in the sequence produced no change to the originalhydrophobic interactions based on the principle of iâ†'i+3and iâ†'i+4hydrophobicinteractions. The single amino acid substitution altered the peptide hydrophobicitydramatically according to RP-HPLC analysis and showed no significant changes onsecondary structure both in aqueous solution and hydrophobic environmentdetermined by circular dichroism. The results showed that the biological activities andhemolytic activities of the analogs exhibited different correlation with hydrophobicity.The biological activities of the peptides needed maintaining hydrophobicity in certaindegree. In this study, increasing the hydrophobicity of the peptides had no effect onthe antibacterial activity, but increased the hemolytic activity dramatically. Whiledecreasing the hydrophobicity of the peptides will decrease the antibacterial activityand the hemolytic activity at the same time. Therefore, peptides hydrophyobicity will be limited in narrow range to optimize the specifity (higher antibacterial activity andlower hemolytic activity). Moreover, the leading peptide V16A showed strongantibacterial activities against Gram-negative and Gram-positive bacteria and weakcytotoxicity against normal mammal cells, thus, may presents a promising moleculefor the development of a peptide antibiotic in clinical practices.Secondly, a cationic α-helical antibacterial peptide HPRP-A1was used as theframework to study the effects of peptide secondary structure on the antimicrobialactivities. We designed a β-sheet peptide and a coiled peptide without changing theoriginal amino acid composition. The secondary structures of peptides weredetermined by CD spectra both in aqueous solution and in hydrophobic environment.The biological activities of peptides were quantified by the minimal concentration ofpeptide against three Gram-negative bacterial strains and three Gram-positivebacterial strains, and the minimal concentration of peptide against human red bloodcells. The results showed that the peptide analogs with increased helicity did notimprove the antibacterial activity, however, their hemolysis activity was enhanced,while the antibacterial activity of peptide analogs with decreased helicity wasoptimized. The peptide with secondary structure transformed to β-sheet showed noimprovement on specificity. The coiled peptide showed dramatic improvement onhemolytic activity.It seemed that helical potency of peptide is correlated more withthe hymolytic activity than with the antibacterial activity in this study.