Interactions of Histidine-Rich Antimicrobial Peptides, Gad-1 and Gad-2, with Model Membranes at low and high pH

Abstract

Antimicrobial peptides (AMPs) are an important component of the innate immune system of many different organisms. Their cationic nature and amphipathic structure make them well suited to interact with and perturb bacterial membranes. A major driving force in the binding of AMPs to bacterial membranes is the electrostatic interaction between positively charged residues of AMPs and anionic lipids of the bacterial membrane. Since histidine has a pKa ∼ 6, histidine-rich AMPs can exhibit greater activity at low pH, when the histidine is positively charged, as compared to at neutral pH, when the histidine is neutral. While Gad-1 and Gad-2 are both histidine-rich AMPs, only Gad-2 exhibits pH-dependent activity against E. coli. Hence, it was surprising to find that neither peptide exhibited pH-dependent membrane disruption in model membranes, when probed by 2H NMR. Thus, we hypothesized the differences in activity might be related to differences in binding affinity. In order to probe the binding, we carried out zeta potential measurements with both Gad-1 and Gad-2 at pH 7.0 and pH 5.0. Additionally, 15N solid state NMR measurements were carried out to establish the topology of the Gad peptides in the bilayer. Together these studies can provide a better understanding of membrane-peptide interactions and the effect of pH on these interactions.