Structure And Alignment Of Membrane-associated Peptaibols By Oriented 15N And 31P Solid-state NMR Spectroscopy

Abstract

Peptaibol antimicrobial peptides are produced by fungi and are characterized by a high content of hydrophobic amino acids, and in particular alpha-isobutyric acid Aib. Here several peptides from this family were uniformly labeled with 15N, purified and reconstituted into oriented phophatidylcholine lipid bilayers and investigated by 15N and 31P solid-state NMR spectroscopy. Whereas alamethicin (20 residues) adopts transmembrane alignments in POPC or DMPC the much shorter ampullosporin A (15) and zervamicin (16) exhibit comparable configurations only in ‘thin’ membranes. In contrast the latter compounds are oriented parallel to the surface in ‘thick’ bilayers indicating that hydrophobic mismatch has a decisive effect. Two-dimensional 15N chemical shift - 1H-15N dipolar coupling solid-state NMR suggests that in their transmembrane configuration ampulosporin and alamethicin adopt mixed alpha-/310-helical structures due to the restraints imposed by the membranes and the bulky Aib residues. The 15N solid-state NMR spectra also provide information on the helical tilt angles, the details of this analysis depend on the appropriate choice of the 15N chemcical shift tensor.Figure: PISEMA spectra of alamethicin (A) and simulations of spectra resulting from 310 (B,C) and mixed 310/α-helical conformations (D).View Large Image | View Hi-Res Image | Download PowerPoint Slide