Advertisement

European Biophysics Journal

, Volume 46, Issue 7, pp 639–646 | Cite as

One pathogen two stones: are Australian tree frog antimicrobial peptides synergistic against human pathogens?

  • Marc-Antoine Sani
  • Siobhan Carne
  • Sarah A. Overall
  • Alexandre Poulhazan
  • Frances Separovic
Original Article

Abstract

Antimicrobial peptides (AMPs) may act by targeting the lipid membranes and disrupting the bilayer structure. In this study, three AMPs from the skin of Australian tree frogs, aurein 1.2, maculatin 1.1 and caerin 1.1, were investigated against Gram-negative Escherichia coli, Gram-positive Staphylococcus aureus, and vesicles that mimic their lipid compositions. Furthermore, equimolar mixtures of the peptides were tested to identify any synergistic interactions in antimicrobial activity. Minimum inhibition concentration and minimum bactericidal concentration assays showed significant activity against S. aureus but not against E. coli. Aurein was the least active while maculatin was the most active peptide and some synergistic effects were observed against S. aureus. Circular dichroism experiments showed that, in the presence of phospholipid vesicles, the peptides transitioned from an unstructured to a predominantly helical conformation (>50%), with greater helicity for POPG/TOCL compared to POPE/POPG vesicles. The helical content, however, was less in the presence of live E. coli and S. aureus, 25 and 5%, respectively. Equimolar concentrations of the peptides did not appear to form greater supramolecular structures. Dye release assays showed that aurein required greater concentration than caerin and maculatin to disrupt the lipid bilayers, and mixtures of the peptides did not cooperate to enhance their lytic activity. Overall, aurein, maculatin, and caerin showed moderate synergy in antimicrobial activity against S. aureus without becoming more structured or enhancement of their membrane-disrupting activity in phospholipid vesicles.

Keywords

Antimicrobial peptides Synergy E. coli S. aureus Phospholipid membrane Circular dichroism 

References

  1. Ambroggio EE, Separovic F, Bowie J, Fidelio GD (2004) Surface behaviour and peptide–lipid interactions of the antibiotic peptides, maculatin and citropin. Biochem Biophys Acta 1664:31–37CrossRefPubMedGoogle Scholar
  2. Apponyi MA, Pukala TL, Brinkworth CS, Maselli VM, Bowie JH, Tyler MJ, Booker GW, Wallace JC, Carver JA, Separovic F, Doyle J, Llewellyn LE (2004) Host-defence peptides of Australian anurans: structure, mechanism of action and evolutionary significance. Peptides 25:1035–1054CrossRefPubMedGoogle Scholar
  3. Blazyk J, Wiegand R, Klein J, Hammer J, Epand RM, Epand RF, Maloy WL, Kari UP (2001) A novel linear amphipathic beta-sheet cationic antimicrobial peptide with enhanced selectivity for bacterial lipids. J Biol Chem 276:27899–27906CrossRefPubMedGoogle Scholar
  4. Cassone M, Otvos L Jr (2010) Synergy among antibacterial peptides and between peptides and small-molecule antibiotics. Expert Rev Anti-infective Ther 8:703–716CrossRefGoogle Scholar
  5. Feng Q, Huang Y, Chen M, Li G, Chen Y (2015) Functional synergy of alpha-helical antimicrobial peptides and traditional antibiotics against Gram-negative and Gram-positive bacteria in vitro and in vivo. Eur J Clin Microbiol Infect Dis 34:197–204CrossRefPubMedGoogle Scholar
  6. Fernandez DI, Le Brun AP, Whitwell TC, Sani MA, James M, Separovic F (2012) The antimicrobial peptide aurein 1.2 disrupts model membranes via the carpet mechanism. Phys Chem Chem Phys 14:15739–15751CrossRefPubMedGoogle Scholar
  7. Fernandez DI, Le Brun AP, Lee TH, Bansal P, Aguilar MI, James M, Separovic F (2013a) Structural effects of the antimicrobial peptide maculatin 1.1 on supported lipid bilayers. Eur Biophys J EBJ 42:47–59CrossRefPubMedGoogle Scholar
  8. Fernandez DI, Lee TH, Sani MA, Aguilar MI, Separovic F (2013b) Proline facilitates membrane insertion of the antimicrobial peptide maculatin 1.1 via surface indentation and subsequent lipid disordering. Biophys J 104:1495–1507CrossRefPubMedPubMedCentralGoogle Scholar
  9. Fernandez DI, Sani MA, Miles AJ, Wallace BA, Separovic F (2013c) Membrane defects enhance the interaction of antimicrobial peptides, aurein 1.2 versus caerin 1.1. Biochem Biophys Acta 1828:1863–1872CrossRefPubMedGoogle Scholar
  10. Hall K, Lee TH, Mechler AI, Swann MJ, Aguilar MI (2014) Real-time measurement of membrane conformational states induced by antimicrobial peptides: balance between recovery and lysis. Sci Rep 4:5479CrossRefPubMedPubMedCentralGoogle Scholar
  11. Hancock RE, Chapple DS (1999) Peptide antibiotics. Antimicrob Agents Chemother 43:1317–1323PubMedPubMedCentralGoogle Scholar
  12. Hancock RE, Lehrer R (1998) Cationic peptides: a new source of antibiotics. Trends Biotechnol 16:82–88CrossRefPubMedGoogle Scholar
  13. Krizsan A, Volke D, Weinert S, Strater N, Knappe D, Hoffmann R (2014) Insect-derived proline-rich antimicrobial peptides kill bacteria by inhibiting bacterial protein translation at the 70S ribosome. Angew Chem 53:12236–12239CrossRefGoogle Scholar
  14. Laadhari M, Arnold AA, Gravel AE, Separovic F, Marcotte I (2016) Interaction of the antimicrobial peptides caerin 1.1 and aurein 1.2 with intact bacteria by 2H solid-state NMR. Biochem Biophys Acta 1858:2959–2964CrossRefPubMedGoogle Scholar
  15. Lee TH, Hall KN, Aguilar MI (2016) Antimicrobial peptide structure and mechanism of action: a focus on the role of membrane structure. Curr Top Med Chem 16:25–39CrossRefPubMedGoogle Scholar
  16. Lobley A, Whitmore L, Wallace BA (2002) DICHROWEB: an interactive website for the analysis of protein secondary structure from circular dichroism spectra. Bioinformatics 18:211–212CrossRefPubMedGoogle Scholar
  17. Nakatsuji T, Gallo RL (2012) Antimicrobial peptides: old molecules with new ideas. J Invest Dermatol 132:887–895CrossRefPubMedGoogle Scholar
  18. Nguyen LT, Haney EF, Vogel HJ (2011) The expanding scope of antimicrobial peptide structures and their modes of action. Trends Biotechnol 29:464–472CrossRefPubMedGoogle Scholar
  19. Nuding S, Frasch T, Schaller M, Stange EF, Zabel LT (2014) Synergistic effects of antimicrobial peptides and antibiotics against Clostridium difficile. Antimicrob Agents Chemother 58:5719–5725CrossRefPubMedPubMedCentralGoogle Scholar
  20. Pasupuleti M, Schmidtchen A, Malmsten M (2012) Antimicrobial peptides: key components of the innate immune system. Crit Rev Biotechnol 32:143–171CrossRefPubMedGoogle Scholar
  21. Rozek T, Waugh RJ, Steinborner ST, Bowie JH, Tyler MJ, Wallace JC (1998) The maculatin peptides from the skin glands of the tree frog Litoria genimaculata: a comparison of the structures and antibacterial activities of maculatin 1.1 and caerin 1.1. J Pept Sci 4:111–115CrossRefPubMedGoogle Scholar
  22. Salditt T, Li C, Spaar A (2006) Structure of antimicrobial peptides and lipid membranes probed by interface-sensitive X-ray scattering. Biochem Biophys Acta 1758:1483–1498CrossRefPubMedGoogle Scholar
  23. Sanchez-Gomez S, Japelj B, Jerala R, Moriyon I, Fernandez Alonso M, Leiva J, Blondelle SE, Andra J, Brandenburg K, Lohner K, Martinez de Tejada G (2011) Structural features governing the activity of lactoferricin-derived peptides that act in synergy with antibiotics against Pseudomonas aeruginosa in vitro and in vivo. Antimicrob Agents Chemother 55:218–228CrossRefPubMedGoogle Scholar
  24. Sani MA, Separovic F (2016) How membrane-active peptides get into lipid membranes. Acc Chem Res 49:1130–1138CrossRefPubMedGoogle Scholar
  25. Sani MA, Whitwell TC, Separovic F (2012) Lipid composition regulates the conformation and insertion of the antimicrobial peptide maculatin 1.1. Biochem Biophys Acta 1818:205–211CrossRefPubMedGoogle Scholar
  26. Sani MA, Whitwell TC, Gehman JD, Robins-Browne RM, Pantarat N, Attard TJ, Reynolds EC, O’Brien-Simpson NM, Separovic F (2013) Maculatin 1.1 disrupts Staphylococcus aureus lipid membranes via a pore mechanism. Antimicrob Agents Chemother 57:3593–3600CrossRefPubMedPubMedCentralGoogle Scholar
  27. Sani MA, Gagne E, Gehman JD, Whitwell TC, Separovic F (2014) Dye-release assay for investigation of antimicrobial peptide activity in a competitive lipid environment. Eur Biophys J EBJ 43:445–450CrossRefPubMedGoogle Scholar
  28. Sani MA, Henriques ST, Weber D, Separovic F (2015a) Bacteria may cope differently from similar membrane damage caused by the Australian tree frog antimicrobial peptide maculatin 1.1. J Biol Chem 290:19853–19862CrossRefPubMedPubMedCentralGoogle Scholar
  29. Sani MA, Lee TH, Aguilar MI, Separovic F (2015b) Proline-15 creates an amphipathic wedge in maculatin 1.1 peptides that drives lipid membrane disruption. Biochem Biophys Acta 1848:2277–2289CrossRefPubMedGoogle Scholar
  30. Shah P, Hsiao FS, Ho YH, Chen CS (2016) The proteome targets of intracellular targeting antimicrobial peptides. Proteomics 16:1225–1237CrossRefPubMedGoogle Scholar
  31. Tachi T, Epand RF, Epand RM, Matsuzaki K (2002) Position-dependent hydrophobicity of the antimicrobial magainin peptide affects the mode of peptide–lipid interactions and selective toxicity. Biochemistry 41:10723–10731CrossRefPubMedGoogle Scholar
  32. Wang Y, Chen CH, Hu D, Ulmschneider MB, Ulmschneider JP (2016) Spontaneous formation of structurally diverse membrane channel architectures from a single antimicrobial peptide. Nature Commun 7:13535CrossRefGoogle Scholar
  33. Whitmore L, Wallace BA (2004) DICHROWEB, an online server for protein secondary structure analyses from circular dichroism spectroscopic data. Nucleic Acids Res 32:W668–W673CrossRefPubMedPubMedCentralGoogle Scholar
  34. Whitmore L, Wallace BA (2008) Protein secondary structure analyses from circular dichroism spectroscopy: methods and reference databases. Biopolymers 89:392–400CrossRefPubMedGoogle Scholar
  35. Zerweck J, Strandberg E, Burck J, Reichert J, Wadhwani P, Kukharenko O, Ulrich AS (2016) Homo- and heteromeric interaction strengths of the synergistic antimicrobial peptides PGLa and magainin 2 in membranes. Eur Biophys J EBJ 45:535–547CrossRefPubMedGoogle Scholar

Copyright information

© European Biophysical Societies' Association 2017

Authors and Affiliations

  1. 1.School of ChemistryBio21 Institute, University of MelbourneMelbourneAustralia
  2. 2.Universite Pierre et Marie Curie (Paris VI)Paris Cedex 5France

Personalised recommendations