The High-Resolution Structures of Two Piscidin Isoforms in Two Microbial Lipid Membrane Mimics Using Solid-State NMR

Piscidins 1 and 3 are twenty-two-residue-long amphipathic, cationic, antimicrobial peptides that have been isolated from hybrid striped bass. Since pathogens are becoming increasingly resistant to traditional antibiotics, antimicrobial peptides such as piscidin may prove to be useful in the future due to their broad-spectrum of activity against microbes and low induction of bacterial resistance. While membrane-active and fast-acting piscidins 1 and 3 have been shown to bind and disrupt bacterial lipid membrane mimics, their mode of action against microbes such as bacteria and yeasts is not known.In earlier studies, we used solid-state NMR to determine the three-dimensional atomic-resolution structures of piscidin 1 and piscidin 3 in aligned bilayers consisting of 3:1 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) and 1,2-dimyristoyl-sn-glycero-3-phospho-rac-glycerol (DMPG). Here, the specific aim has been to solve the structures of these peptides in 1:1 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) to 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) so that we could compare and contrast these structures with those in 3:1 DMPC/DMPG, a lipid system that has lower anionic content and does not exhibit the negative intrinsic membrane curvature of bacterial membranes. We have collected two-dimensional solid-state NMR spectra on 15N-backbone labeled piscidins in the presence of aligned 1:1 POPE/POPG bilayers. 15N chemical shifts and 15N-1H dipolar couplings have been combined to obtain the topological orientation of the peptides and calculate their atomic level backbone structures in the membrane-bound state. The data indicate that the carboxyl- and amino-ends of the peptides are affected differently by the changes in lipid composition. We will discuss the implications in terms of structural features and peptide-lipid interactions that may be important for piscidin's mode of action. Overall, these studies performed using native-like conditions provide us with a better understanding of structure-function relationships in piscidins and related antimicrobial peptides.