Abstract
Maculatin 1.1 (Mac1) is an antimicrobial peptide (AMP) from the skin secretions of Australian tree frogs. In this work, the interaction of Mac1 with anionic phospholipid bilayers was investigated by NMR, circular dichroism (CD) spectroscopy, neutron reflectometry (NR) and molecular dynamics (MD). In buffer, the peptide is unstructured but in the presence of anionic (DPC/LMPG) micelles or (DMPC/DMPG/DHPC) bicelles adopts a helical structure. Addition of the soluble paramagnetic agent gadolinium (Gd-DTPA) into the Mac1-DPC/LMPG micelle solution showed that the N-terminus is more exposed to the hydrophilic Gd-DTPA than the C-terminus in micelles. 2H and 31P solid-state NMR showed that Mac1 had a greater effect on the anionic lipid (DMPG). A deuterium labeled Mac1 used in NR experiments indicated that the AMP spanned across anionic (PC/PG) bilayers, which was compatible with MD simulations. Simulations also showed that Mac1 orientation remained transmembrane in bilayers and wrapped on the surface of the micelles regardless of the lipid or detergent charge. Thus, the peptide orientation appears to be more susceptible to curvature than charged surface. These results support the formation of transmembrane pores by Mac1 in model bacterial membranes.
Highlights
Maculatin 1.1 (Mac1)s is an antimicrobial peptide (AMP) from the skin secretions of the Australian tree frog Litoria genimaculata (Rozek et al, 1998)
Mac1 will bind to a range of different model membranes that replicate various characteristics of the Gram-positive bacterial membrane (Sani et al, 2015a), and in this study we investigate the orientation of Mac1 in anionic phosphatidylglycerol (PG) membranes that better reflect the charge state of bacterial membranes which is the principal target of Mac1
Comparing the 15N HSQC obtained in DPC micelles and DPC/LMPG (9:1) showed that the N-terminus of Mac1 exhibited small chemical shift changes at Val5, Leu6, and Ala7 while the middle and C-terminus sections exhibited more pronounced chemical shift perturbations, the highest observed for Ala16 (Figure 1A)
Summary
Maculatin 1.1 (Mac1)s is an antimicrobial peptide (AMP) from the skin secretions of the Australian tree frog Litoria genimaculata (Rozek et al, 1998). The peptide forms part of the frog’s innate immune system and is effective at killing a wide range of Gram-positive bacteria (Fernandez et al, 2009) This makes Mac, along with a range of other AMPs found across nature, a possibility for development of alternative antibiotics (Lee et al, 2015). The final characteristic to note is that Mac has a proline residue that induces a kink in the structure of the peptide, and substitution of this residue with alanine or glycine results in reduced binding to membranes (Fernandez et al, 2013b) This is the result of the proline creating a wedge which allows the peptide to insert into the bilayer (Sani et al, 2015b)
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