Skip to main content
SpringerLink
Log in

Lipid interactions of LAH4, a peptide with antimicrobial and nucleic acid transfection activities

  • Original Paper
  • Published:
European Biophysics Journal Aims and scope Submit manuscript

Abstract

The cationic amphipathic designer peptide LAH4 exhibits potent antimicrobial, nucleic acid transfection and cell penetration activities. Closely related derivatives have been developed to enhance viral transduction for gene therapeutic assays. LAH4 contains four histidines and, consequently, its overall charge and membrane topology in lipid bilayers are strongly pH dependent. In order to better understand the differential interactions of this amphipathic peptide with negatively-charged membranes its interactions, topologies, and penetration depth were investigated in the presence of lipid bilayers as a function of pH, buffer, phospholipid head group, and fatty acyl chain composition using a combination of oriented synchrotron radiation circular dichroism spectroscopy as well as oriented and non-oriented solid-state NMR spectroscopy. This combination of methods indicates that in the presence of lipids with phosphatidylglycerol head groups, the topological equilibria of LAH4 is shifted towards more in-plane configurations even at neutral pH. In contrast, a transmembrane alignment is promoted when LAH4 interacts with membranes made of dimyristoyl phospholipids rather than palmitoyl-oleoyl-phospholipids. Finally, the addition of citrate buffer favours LAH4 transmembrane alignments, even at low pH, probably by complex formation with the cationic charges of the peptide. In summary, this study has revealed that the membrane topology of this peptide is readily modulated by the environmental conditions.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

Abbreviations

CD:

Circular dichroism

DMPC:

1, 2-Dimyristoyl-sn-glycero-3-phosphocholine

DMPG:

1, 2-Dimyristoyl-sn-glycero-3-phospho-(1'-rac-glycerol)

DSPC:

1, 2-Distearoyl-sn-glycero-3-phosphocholine

DPPC:

1, 2-Dipalmitoyl-sn-glycero-3-phosphocholine

hΦ17, KALP:

GKKLA LALAL ALALA LALAL KKA-CONH2

KL14:

KKLLK KAKKL LKKL-CONH2

LAH4:

KKALL ALALH HLAHL ALHLA LALKK A-CONH2

NMR:

Nuclear magnetic resonance

oSRCD:

Oriented synchrotron radiation circular dichroism

PC:

Phosphatidylcholine

PG:

Phosphatidylglycerol

PGLa:

GMASK AGAIA GKIAK VALKA L-CONH2

POPC:

1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine

POPG:

1-Palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1'-rac-glycerol)

r.h.:

Relative humidity

SAXS:

Small angle X-ray scattering

SRCD:

Synchrotron radiation circular dichroism

References

  • Aisenbrey C, Goormaghtigh E, Ruysschaert JM, Bechinger B (2006a) Translocation of amino acyl residues from the membrane interface to the hydrophobic core: thermodynamic model and experimental analysis using ATR-FTIR spectroscopy. Mol Membr Biol 23:363–374

    Article  PubMed  CAS  Google Scholar 

  • Aisenbrey C, Kinder R, Goormaghtigh E, Ruysschaert JM, Bechinger B (2006b) Interactions involved in the realignment of membrane-associated helices: an investigation using oriented solid-state NMR and ATR-FTIR spectroscopies topologies. J Biol Chem 281:7708–7716

    Article  PubMed  CAS  Google Scholar 

  • Aisenbrey C, Sudheendra US, Ridley H, Bertani P, Marquette A, Nedelkina S, Lakey JH, Bechinger B (2007) Helix orientations in membrane-associated Bcl-X L determined by 15N solid-state NMR spectroscopy. Eur Biophys J 36:451–460

    Article  PubMed  CAS  Google Scholar 

  • Aisenbrey C, Michalek M, Salnikov ES, Bechinger B (2013) Solid-state NMR approaches to study protein structure and protein-lipid interactions. In: Kleinschmidt JH (ed) Lipid-protein interactions: methods and protocols. Springer, New York, pp 357–387

    Chapter  Google Scholar 

  • Bechinger B (1996) Towards membrane protein design:pH dependent topology of histidine-containing polypeptides. J Mol Biol 263:768–775

    Article  PubMed  CAS  Google Scholar 

  • Bechinger B (1999) The structure, dynamics and orientation of antimicrobial peptides in membranes by solid-state NMR spectroscopy. Biochim Biophys Acta 1462:157–183

    Article  PubMed  CAS  Google Scholar 

  • Bechinger B (2009) Rationalizing the membrane interactions of cationic amphipathic antimicrobial peptides by their molecular shape. Cur Opin Colloid Interface Sci Surfactants 14:349–355

    Article  CAS  Google Scholar 

  • Bechinger B (2011) Insights into the mechanisms of action of host defence peptides from biophysical and structural investigations. J Pept Sci 17:306–314

    Article  PubMed  CAS  Google Scholar 

  • Bechinger B, Sizun C (2003) Alignment and structural analysis of membrane polypeptides by 15N and 31P solid-state NMR spectroscopy. Concepts Magn Reson 18A:130–145

    Article  CAS  Google Scholar 

  • Bechinger B, Ruysschaert JM, Goormaghtigh E (1999) Membrane helix orientation from linear dichroism of infrared attenuated total reflection spectra. Biophys J 76:552–563

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Bechinger B, Bertani P, Werten S, Mendonca de Moraes C, Aisenbrey C, Mason AJ, Perrone B, Prudhon M, Sudheendra US, Vidovic V (2010) The structural and topological analysis of membrane polypeptides by oriented solid-state NMR spectroscopy: sample preparation and theory. In: Miguel C (ed) Membrane-active peptides: methods and results on structure and function. International University Line, La Jolla, pp 196–215

    Google Scholar 

  • Burck J, Roth S, Wadhwani P, Afonin S, Kanithasen N, Strandberg E, Ulrich AS (2008) Conformation and membrane orientation of amphiphilic helical peptides by oriented circular dichroism. Biophys J 95:3872–3881

    Article  PubMed  PubMed Central  Google Scholar 

  • Cheng JTJ, Hale JD, Elliott M, Hancock REW, Straus SK (2011) The importance of bacterial membrane composition in the structure and function of aurein 2.2 and selected variants. Biochim Biophys Acta 1808:622–633

    Article  PubMed  CAS  Google Scholar 

  • Fenard D, Ingrao D, Seye A, Buisset J, Genries S, Martin S, Kichler A, Galy A (2013) Vectofusins, a new class of viral entry enhancers, strongly promote lentiviral transduction of human hematopoietic stem cells. Mol Ther Nucleic Acids 2:e90. doi:10.1038/mtna.2013.1017

    Article  PubMed  Google Scholar 

  • Gemmill KB, Muttenthaler M, Delehanty JB, Stewart MH, Susumu K, Dawson PE, Medintz IL (2013) Evaluation of diverse peptidyl motifs for cellular delivery of semiconductor quantum dots. Anal Bioanal Chem 405:6145–6154

    Article  PubMed  CAS  Google Scholar 

  • Georgescu J, Verly RM, Bechinger B (2010) NMR structures of the histidine-rich peptide LAH4 in micellar environments: membrane insertion, pH-dependent mode of antimicrobial action and DNA transfection. Biophys J 99:2507–2515

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Goldmann WH, Bechinger B, Lele T (2008) Cytoskeletal proteins at the lipid membrane. In:Tien HT, Ottova-Leitmannova A (eds) Planar lipid bilayers and their applications:recent progress. vol 6, Elsevier Inc., pp 227–255

  • Harzer U, Bechinger B (2000) The alignment of lysine-anchored membrane peptides under conditions of hydrophobic mismatch: a CD, 15N and 31P solid-state NMR spectroscopy investigation. Biochem 39:13106–13114

    Article  CAS  Google Scholar 

  • He K, Ludtke SJ, Heller WT, Huang HW (1996) Mechnaism of alamethicin insertion into lipid bilayers. Biophys J 71:2669–2679

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Hediger S, Meier BH, Kurur ND, Bodenhausen G, Ernst RR (1994) NMR cross-polarization by adiabatic passage through the Hartmann-Hahn condition (APHH). Chem Phys Lett 223:283–288

    Article  CAS  Google Scholar 

  • Hirschinger J, Raya J, Perrone B, Bechinger B (2011) Chemical shift powder spectra obtained by using Rotor-directed exchange of orientations cross-polarization (RODEO-CP). Chem Phys Lett 508:155–164

    Article  Google Scholar 

  • Huang HW (2000) Action of antimicrobial peptides: two-state model. Biochem 39:8347–8352

    Article  CAS  Google Scholar 

  • Israelachvili JN, Marcelja S, Horn RG (1980) Physical principles of membrane organization. Q Rev Biophys 13:121–200

    Article  PubMed  CAS  Google Scholar 

  • Kemayo Koumkoua P, Aisenbrey C, Salnikov ES, Rifi O, Bechinger B (2014) On the design of supramolecular assemblies made of peptides and lipid bilayers. J Pept Sci 20(7):526–536

    Article  PubMed  CAS  Google Scholar 

  • Kichler A, Leborgne C, März J, Danos O, Bechinger B (2003) Histidine-rich amphipathic peptide antibiotics promote efficient delivery of DNA into mammalian cells. Proc Natl Acad Sci USA 100:1564–1568

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Killian JA (2003) Synthetic peptides as models for intrinsic membrane proteins. FEBS Lett 555:134–138

    Article  PubMed  CAS  Google Scholar 

  • Kollmitzer B, Heftberger P, Rappolt M, Pabst G (2013) Monolayer spontaneous curvature of raft-forming membrane lipids. Soft Matter 9:10877–10884

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Langlet-Bertin B, Leborgne C, Scherman D, Bechinger B, Mason AJ, Kichler A (2010) Design and evaluation of histidine-rich amphipathic peptides for siRNA delivery. Pharm Res 27:1426–1436

    Article  PubMed  CAS  Google Scholar 

  • Liu Y, Kim YJ, Ji M, Fang J, Siriwon N, Zhang L, Wang P (2014) Enhancing gene delivery of adeno-associated viruses by cell-permeable peptides. Mol Thera—Method Clin Dev 1. doi:10.1038/mtm.2013.1012

  • Marquette A, Mason AJ, Bechinger B (2008) Aggregation and membrane permeabilizing properties of designed histidine-containing cationic linear peptide antibiotics. J Pept Sci 14:488–495

    Article  PubMed  CAS  Google Scholar 

  • Mason AJ, Gasnier C, Kichler A, Prevost G, Aunis D, Metz-Boutigue MH, Bechinger B (2006a) Enhanced membrane disruption and antibiotic action against pathogenic bacteria by designed histidine-rich peptides at acidic pH. Antimicrob Agents Chemother 50:3305–3311

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Mason AJ, Husnal-Chotimah IN, Bertani P, Bechinger B (2006b) A spectroscopic study of the membrane interaction of the antimicrobial peptide pleurocidin. Mol Membr Biol 23:185–194

    Article  PubMed  CAS  Google Scholar 

  • Mason AJ, Martinez A, Glaubitz C, Danos O, Kichler A, Bechinger B (2006c) The antibiotic and DNA-transfecting peptide LAH4 selectively associates with, and disorders, anionic lipids in mixed membranes. FASEB J 20:320–322

    PubMed  CAS  Google Scholar 

  • Mason AJ, Bertani P, Moulay G, Marquette A, Perrone B, Drake AF, Kichler A, Bechinger B (2007a) Membrane interaction of chrysophsin-1, a histidine-rich antimicrobial peptide from red sea bream. Biochem 46:15175–15187

    Article  CAS  Google Scholar 

  • Mason AJ, Leborgne C, Moulay G, Martinez A, Danos O, Bechinger B, Kichler A (2007b) Optimising histidine rich peptides for efficient DNA delivery in the presence of serum. J Control Release 118:95–104

    Article  PubMed  CAS  Google Scholar 

  • Mason AJ, Moussaoui W, Abdelrhaman T, Boukhari A, Bertani P, Marquette A, Shooshtarizaheh P, Moulay G, Boehm N, Guerold B, Sawers RJH, Kichler A, Metz-Boutigue MH, Candolfi E, Prevost G, Bechinger B (2009) Structural determinants of antimicrobial and antiplasmodial activity and selectivity in histidine rich amphipathic cationic peptides. J Biol Chem 284:119–133

    Article  PubMed  CAS  Google Scholar 

  • Matsuzaki K, Harada M, Funakoshi S, Fujii N, Miyajima K (1991) Physicochemical determinants for the interactions of magainins 1 and 2 with acidic lipid bilayers. Biochim Biophys Acta 1063:162–170

    Article  PubMed  CAS  Google Scholar 

  • Matsuzaki K, Sugishita K, Ishibe N, Ueha M, Nakata S, Miyajima K, Epand RM (1998) Relationship of membrane curvature to the formation of pores by magainin 2. Biochem 37:11856–11863

    Article  CAS  Google Scholar 

  • Prongidi-Fix L, Bertani P, Bechinger B (2007a) The membrane alignment of helical peptides from non-oriented 15N chemical shift solid-state NMR spectroscopy. J Am Chem Soc 129:8430–8431

    Article  PubMed  CAS  Google Scholar 

  • Prongidi-Fix L, Sugewara M, Bertani P, Raya J, Leborgne C, Kichler A, Bechinger B (2007b) Self-promoted uptake of peptide/DNA transfection complexes. Biochem 46:11253–11262

    Article  CAS  Google Scholar 

  • Salnikov E, Bechinger B (2011) Lipid-mediated peptide–peptide interactions in bilayers: structural insights into the synergistic enhancement of the antimicrobial activities of PGLa and magainin 2. Biophys J 100:1473–1480

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Salnikov ES, Mason AJ, Bechinger B (2009) Membrane order perturbation in the presence of antimicrobial peptides by 2H solid-state NMR spectroscopy. Biochimie 91:743

    Article  Google Scholar 

  • Salnikov E, Aisenbrey C, Vidovic V, Bechinger B (2010) Solid-state NMR approaches to measure topological equilibria and dynamics of membrane polypeptides. Biochim Biophys Acta 1798:258–265

    Article  PubMed  CAS  Google Scholar 

  • Sansom MS (1993) Alamethicin and related peptaibols–model ion channels. Eur Biophys J 22:105–124

    PubMed  CAS  Google Scholar 

  • Strandberg E, Tiltak D, Ehni S, Wadhwani P, Ulrich AS (2012) Lipid shape is a key factor for membrane interactions of amphipathic helical peptides. Biochim Biophys Acta 1818:1764–1776

    Article  PubMed  CAS  Google Scholar 

  • Strandberg E, Zerweck J, Wadhwani P, Ulrich AS (2013) Synergistic insertion of antimicrobial magainin-family peptides in membranes depends on the lipid spontaneous curvature. Biophys J 104:L09–L11

    Google Scholar 

  • Vogel H (1987) Comparison of the conformation and orientation of alamethicin and melittin in lipid membranes. Biochem 26:4562–4572

    Article  CAS  Google Scholar 

  • Vogt TCB, Bechinger B (1999) The interactions of histidine-containing amphipathic helical peptide antibiotics with lipid bilayers: the effects of charges and pH. J Biol Chem 274:29115–29121

    Article  PubMed  CAS  Google Scholar 

  • White SH, Wimley WC (1999) Membrane protein folding and stability: physical principles. Annu Rev Biophys Biomol Struct 28:319–365

    Article  PubMed  CAS  Google Scholar 

  • Wien F, Wallace BA (2005) Calcium fluoride micro cells for synchrotron radiation circular dichroism spectroscopy. Appl Spectrosc 59:1109–1113

    Article  PubMed  CAS  Google Scholar 

  • Woolley GA, Wallace BA (1992) Model ion channels: gramicidin and alamethicin. J Membr Biol 129:109–136

    PubMed  CAS  Google Scholar 

  • Wu Y, Huang HW, Olah GA (1990) Method of oriented circular dichroism. Biophys J 57:797–806

    Article  PubMed  CAS  PubMed Central  Google Scholar 

  • Zasloff M (2002) Antimicrobial peptides of multicellular organisms. Nature 415:389–395

    Article  PubMed  CAS  Google Scholar 

  • Zhang TT, Khang TH, Ma B, Xu Y, Hung CF, Wu TC (2011) LAH4 enhances CD8+ T cell immunity of protein/peptide-based vaccines. Vaccine 30:784–793

    Article  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgments

BP was supported by a studentship from the European Commission under the 6th Framework Programme through the Marie-Curie Action: BIOCONTROL, contract number MCRTN–33439. Financial contributions (to BB) by the Agence Nationale de la Recherche (projects TRANSPEP, MembraneDNP, and the LabEx Chemistry of Complex Systems), the University of Strasbourg, the CNRS, the Région Alsace, the RTRA International Center of Frontier Research in Chemistry and the French Foundation for Medical Research (FRM) are gratefully acknowledged. This project was also supported by grants from the UK Biotechnology and Biological Sciences Research Council (to BAW) and an SRCD beamtime grant (to BAW) from the ISA (Aarhus, Denmark) Synchrotron, which is supported by the EU Integrated Infrastructure Initiative (I3), European Light Sources Activities (ELISA), under grant agreement No 226716.

Author information

Authors and Affiliations

  1. UMR7177, Institut de Chimie, Université de Strasbourg/CNRS, 1, rue Blaise Pascal, Strasbourg, 67070, France

    Barbara Perrone, Evgeniy S. Salnikov & Burkhard Bechinger

  2. Institute of Structural and Molecular Biology, Birkbeck College, University of London, Malet Street, London, WC1E 7HX, UK

    Andrew J. Miles & B. A. Wallace

Authors
  1. Barbara Perrone
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Andrew J. Miles
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Evgeniy S. Salnikov
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. B. A. Wallace
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Burkhard Bechinger
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to B. A. Wallace or Burkhard Bechinger.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Perrone, B., Miles, A.J., Salnikov, E.S. et al. Lipid interactions of LAH4, a peptide with antimicrobial and nucleic acid transfection activities. Eur Biophys J 43, 499–507 (2014). https://doi.org/10.1007/s00249-014-0980-y

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00249-014-0980-y

Keywords

Search

Navigation