High-Resolution Structure of Piscidin in Aligned Lipid Bilayers: Implications for Antimicrobial Mode of Action

Abstract

Piscidin is an amphipathic cationic antimicrobial peptide that belongs to a large family of versatile host-defense peptides, which interact, at least initially, with cell membranes in order to perform their function. In the research presented here, we characterize the secondary structure and dynamics of piscidin in order to identify factors optimizing specific molecular interactions that are directly related to its function and mode of action. Our long term goal is to identify common principles that will facilitate the design of pharmaceuticals with broad-spectrum antibacterial activity, minimum induction of bacterial resistance, and low toxicity for mammalian cells.Previously, we demonstrated that membrane-bound piscidin 1 (p1) adopts an alpha-helical structure, which lies in the plane of the bilayer and experiences fast motions. Here, high-resolution solid-state NMR spectra have been obtained from multiply 15N-backbone-labeled p1 aligned in hydrated lipid bilayers. Analysis of data from twenty sites of this 22-mer reveals two helical segments separated by a kink at Gly13. This kink may help one portion of the peptide insert more deeply in the hydrophobic lipid bilayer, which may be related to the mechanism of membrane disruption. To characterize water exposure, hydrogen-deuterium exchange experiments have been performed on 15N-backbone labeled samples. In addition, solid-state NMR was applied to 15N-side-chain-labeled His-17 p1 to titrate this side chain, which resides at the interface between the hydrophilic and hydrophobic domains of p1. Overall, our atomic-level investigation of the structure, dynamics, and water exposure of membrane-bound piscidin provides new insights into its mode of action and thus help understand how antimicrobial peptides recognize membranes and initiate their activities on microbial cells.