ORIGINAL PAPER Investigations into the ability of the peptide, HAL18, to interact with bacterial membranes Sarah R. Dennison Æ Young Soo Kim Æ Hyung Joon Cha Æ David A. Phoenix Received: 7 April 2008 / Revised: 6 June 2008 / Accepted: 12 June 2008 / Published online: 4 July 2008 Ó European Biophysical Societies’ Association 2008 Abstract Halocidin was isolated from hemocytes, Halo- cynthia aurantium as a heterodimeric peptide consisting of two a-helical subunits, Hal15 and Hal18. Hal18 was shown to have antibacterial properties against Bacillus subtilis (MLC = 15 lM) and Escherichia coli (MLC = 100 lM). The peptide was shown to produce stable monolayers, which were characteristic of a-helical peptides predicted to orientate parallel to the surface of the interface. Constant area assays showed that Hal18 was surface active (4 lM) inducing surface pressure changes [ 30 mN m -1 charac- teristic of membrane interactive peptides. The peptide induced stable surface pressure changes in monolayers that were mimetic of B. subtilis membranes (circa 7 mN m -1 ) and E. coli membrane-mimics (circa 4 mN m -1 ). Hal18 inserted readily into zwitterionic DOPE and anionic DOPG monolayers inducing surface pressure changes circa 8 mN m -1 in both cases, providing evidence that interac- tion is not headgroup specific. Thermodynamic analysis of compression isotherms showed that the presence of Hal18 destabilised B. subtilis membranes (DG Mix [ 0), which is in contrast to its stabilising effect on E. coli lipid extract implying the differential antimicrobial efficacy may be driven by lipid packing. Keywords Antimicrobial peptide  Monolayer stability  Peptide monolayer  Lipid–peptide interactions  Thermodynamic analysis Abbreviations a Interaction parameter DH Mixing enthalpy CL Cardiolipin C s -1 Compressibility modulus DOPE Dioleoylphosphatidylethanolamine DOPG Dioleoylphosphatidylglycerol a-AMPs Alpha helical antimicrobial peptides DG Mix Gibbs free energy of mixing Introduction In recent years overuse of antibiotics has led to the emer- gence and spread of drug-resistant pathogens. For example, there are now widespread reports of methacillin-resistant Staphylococcus aureus (MRSA), and bacterium that are no longer sensitive to antibiotics of last resort such as vancomycin (Hiramatsu et al. 1997). The pharmaceutical industry is therefore, constantly searching for new and effective antimicrobial agents, especially for the treatment of multiple drug-resistant bacteria (Diamond 2001; Reddy et al. 2004; Schroder and Harder 2006; Toke 2005; Zasloff 2002). In recent years considerable attention has been given to a-helical antimicrobial peptides (a-AMPs) that exhibit broad spectrum antimicrobial activity (Boman 1995; Zasloff 2002) against Gram-positive bacteria, Gram- negative bacteria, and fungi. These agents seem to have limited susceptibility to drug resistance yet despite inten- sive study there remains a lack of understanding of their structure/function relationships. a-AMPs may utilise multiple modes of action. Those with even hydrophobicity distribution along the helical long axis have been postulated to support a mode of action S. R. Dennison  D. A. Phoenix (&) Faculty of Science and Technology, University of Central Lancashire, Preston PR1 2HE, UK e-mail: daphoenix@uclan.ac.uk Y. S. Kim  H. J. Cha Department of Chemical Engineering, Pohang University of Science and Technology, Pohang 790-784, Korea 123 Eur Biophys J (2008) 38:37–43 DOI 10.1007/s00249-008-0352-6