Trichogin GA IV Alignment and Oligomerization in Phospholipid Bilayers

Abstract

Trichogin GA IV is a short peptaibol with antimicrobial activity. This uncharged, but amphipathic, sequence is aligned at the membrane interface and undergoes a transition to an aggregated state that inserts more deeply into the membrane, an assembly that predominates at a peptide-to-lipid ratio (P/L) of 1:20. In this work, the natural trichogin sequence was prepared and reconstituted into oriented lipid bilayers. The ¹⁵ N NMR chemical shift is indicative of a well-defined alignment of the peptide parallel to the membrane surface at P/Ls of 1:120 and 1:20. When the P/L is increased to 1:8, an additional peptide topology is observed that is indicative of a heterogeneous orientation, with helix alignments ranging from around the magic angle to perfectly in-plane. The topological preference of the trichogin helix for an orientation parallel to the membrane surface was confirmed by attenuated total reflection FTIR spectroscopy. Furthermore, ¹⁹ F CODEX experiments were performed on a trichogin sequence with ¹⁹ F-Phe at position 10. The CODEX decay is in agreement with a tetrameric complex, in which the ¹⁹ F sites are about 9-9.5 Å apart. Thus, a model emerges in which the monomeric peptide aligns along the membrane surface. When the peptide concentration increases, first dimeric and then tetrameric assemblies form, made up from helices oriented predominantly parallel to the membrane surface. The formation of these aggregates correlates with the release of vesicle contents including relatively large molecules.

Keywords: ATR FTIR spectroscopy; antimicrobial peptides; lipid bilayers; membrane permeabilization; solid-state NMR spectroscopy.