While
antimicrobial peptides (AMPs) have been widely investigated
as potential therapeutics, high-resolution structures obtained under
biologically relevant conditions are lacking. Here, the high-resolution
structures of the homologous 22-residue long AMPs piscidin 1 (p1)
and piscidin 3 (p3) are determined in fluid-phase 3:1 phosphatidylcholine/phosphatidylglycerol
(PC/PG) and 1:1 phosphatidylethanolamine/phosphatidylglycerol (PE/PG)
bilayers to identify molecular features important for membrane destabilization
in bacterial cell membrane mimics. Structural refinement of 1H–15N dipolar couplings and 15N chemical
shifts measured by oriented sample solid-state NMR and all-atom molecular
dynamics (MD) simulations provide structural and orientational information
of high precision and accuracy about these interfacially bound α-helical
peptides. The tilt of the helical axis, τ, is between 83°
and 93° with respect to the bilayer normal for all systems and
analysis methods. The average azimuthal rotation, ρ, is 235°,
which results in burial of hydrophobic residues in the bilayer. The
refined NMR and MD structures reveal a slight kink at G13 that delineates
two helical segments characterized by a small difference in their
τ angles (<10°) and significant difference in their
ρ angles (∼25°). Remarkably, the kink, at the end
of a G(X)4G motif highly conserved among members of the
piscidin family, allows p1 and p3 to adopt ρ angles that maximize
their hydrophobic moments. Two structural features differentiate the
more potent p1 from p3: p1 has a larger ρ angle and less N-terminal
fraying. The peptides have comparable depths of insertion in PC/PG,
but p3 is 1.2 Å more deeply inserted than p1 in PE/PG. In contrast
to the ideal α-helical structures typically assumed in mechanistic
models of AMPs, p1 and p3 adopt disrupted α-helical backbones
that correct for differences in the amphipathicity of their N- and
C-ends, and their centers of mass lie ∼1.2–3.6 Å
below the plane defined by the C2 atoms of the lipid acyl chains.
P and 2 H NMR in mechanically aligned bilayers composed of a neutral membrane with a zwitterionic head group (DMPC) and a bacterial-like membrane composed of a 3:1 mixture of neutral (DMPC) and anionic lipids (DMPG). 2 H NMR spectra suggest weaker binding or less well-defined orientations of the Retro-LfB6 peptides compared to native LfB6 peptides. 31 P NMR spectra reveal that the lipids remain primarily in a bilayer arrangement, with the peptides causing little change to the phosphate head groups. Antimicrobial assays demonstrate that the C6-acylated, Trp-methylated, Retro-LfB6 peptide has enhanced activity relative to the native peptide against S. aureus. Results from partitioning assays will be compared with the NMR and antimicrobial data.
The antimicrobial peptides (AMPs) PGLa and magainin 2 (MAG2) found in the skin of the African frog Xenopus laevis show a synergistic enhancement of their activity [1]. Both peptides form amphipathic a-helices when binding to a lipid membrane, and their orientation in membranes have been determined with high accuracy using solid state 2 H-, 15 N-, and 19 F-NMR. We have previously shown that PGLa inserts into DMPC/DMPG bilayers in the presence of an equimolar amount of MAG2, but not on its own, even at high concentrations [2]. This indicates formation of stable heterodimeric peptide pores, which in turn can explain the synergism between the peptides. Here, we show that MAG2, both with and without PGLa, always stays almost flat on the membrane surface; however, there is a small change in the orientation in the presence of PGLa, and dynamics is reduced, indicating that a PGLa-MAG2 complex is formed. In POPC/POPG, both peptides stay flat on the membrane surface, alone or combined. This is in agreement with our previous finding that the AMP MSI-103 always stays flat on the surface in unsaturated lipids (or more generally in lipid systems with negative spontaneous curvature), but can insert deeper into the membrane in saturated lipids (where the spontaneous curvature is positive) [3].
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.