Charge Density as a Determinant of Antimicrobial Activity of Small Cationic Peptides

Abstract

Purpose: : A series of cationic peptides were synthesized to examine effect of charge density on antimicrobial activity. The structural parameters to antibacterial activity of analogues were studied by ¹H NMR and molecular dynamics (MD)

Methods: : Eight C-terminus analogues of hBD3 [W2, F2, Y2, V2, L2, I2 and H2, C2] were synthesized by SPPS. The antibacterial activity of peptides against P. aeruginosa was manifested as the logarithmic reduction of bacteria. Cytotoxicity of analogues was analyzed on human conjunctiva epithelial cells by measuring cell viability using CellTiter-blue. HBD3 was used as a control. The structures of Y2, F2 and C2 were determined by NMR and MD.

Results: : W2, V2, and especially Y2, showed high activity selectively against P. aeruginosa at 4.3-9.7 µM. No cytotoxicity of this series of analogues was observed even at concentrations up to 200 µg/mL. The physico-chemical properties distinguishing active Y2 and less active F2 and C2 include propensities to oligomerize and to accrete unique structure both in water and on the surface of lipid membranes. MD showed a higher stability of Y2- and F2-dimers compared to C2. The prominent structural difference between Y2 and F2 dimers might underline differences in their activity. Y2 dimer maintains a hairpin like, more compact configuration, in striking contrast to a twisted helical, extended structure of F2 dimer. The compact structure of Y2 induces a higher charge density. The results show that the binding affinity and partitioning into the lipid phase, the ability to dimerize and accrete well defined structures upon interactions with lipid membranes contribute to compactization of positive charges within peptide oligomers.

Conclusions: : The charge density of peptide was found to directly correlate with antimicrobial activity. These novel observations may provide a new basis for design of improved antimicrobial peptides.

Keywords: protein structure/function • antibiotics/antifungals/antiparasitics