Characterization of Nucleic Acid Binding by the Histone-Derived Antimicrobial Peptides Buforin II and DesHDAP1

Abstract

Antimicrobial peptides (AMPs), which are found in numerous living organisms, are active against a wide range of bacteria and other pathogens, making them promising candidates for alternatives to conventional antibiotics. While many AMPs inhibit bacterial growth through membrane disruption, some AMPs, including buforin II and DesHDAP1, are hypothesized to kill bacteria by binding to intracellular nucleic acids after translocating across the bacterial cell membrane without significant membrane permeabilization. To understand this lesser known mechanism, the peptide-nucleic acid binding interactions of these systems were investigated using several experimental and computational methods. The peptide-nucleic acid binding for buforin II and DesHDAP1 were measured experimentally using a fluorescence intercalator displacement (FID) assay. In these experiments, having an increased composition of basic residues that are arginine versus lysine were shown to promote binding for variants of both buforin II and DesHDAP1. When binding was tested using different combinations of DNA sequences, neither buforin II nor DesHDAP1 significantly favored particular DNA sequences. To provide structural explanations for experimental results, molecular dynamics (MD) simulations, pKa calculations, electrostatics calculations via component analysis were used. Using these methods, particular arginine residues within each peptide were found to interact more favorably with DNA. Results from MD simulations also showed that nucleic acid-peptide binding was mostly due to interactions between the peptide and phosphate backbone of nucleic acids, providing an explanation for the lack of sequence specificity observed experimentally. These insights regarding nucleic acid binding of buforin II and DesHDAP1, paired with a deeper understanding of the peptides’ structures and membrane interactions, are necessary for development of novel pharmaceutical applications using AMPs.