Melittin

The POPC (1-palmitoyl-2-oleoyl-glycero-3-phosphocholine) was purchased from Avanti Polar Lipids (Birmingham, USA) through Sigma–Aldrich (Castle Hill, Australia). Mechanical extrusion for the preparation of lipid vesicles was carried out using an extruder from Avanti Polar Lipids (Birmingham, USA). Melittin was purchased from AnaSpec (Fremont, USA). Reagents for buffer solutions, glycine-HCl and CHAPS (3-((3-cholamidopropyl) dimethylammonio)-1-propanesulfonate) solutions were purchased from Sigma–Aldrich (Castle Hill, Australia). Running buffer for SPR experiments consisted of 0.05 M HEPES (Sigma–Aldrich, Australia) at a pH of 7.4 and also included 150 mM NaCl for experiments investigating the effect of ionic strength. The SPR experiments were conducted in a BiacoreT200 instrument using a L1 sensor chip S series (GE Healthcare Life Sciences, Paramatta, Australia).

Lyophilized peptides LL37 (LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPRTES), melittin (GIGAVLKVLTTGLPALISWIKRKRQQ), buforin (TRSSRAGLQFPVGRVHRLLRK), MM1 (GIGAVLKALTTGLGALASAIKRKRQQ), AR23 (AIGSILGALAKGLPTLISWIKNR), and RV23 (RIGVLLARLPKLFSLFKLMGKKV) were purchased from Anaspec or Lifetein (≥95% purity by high-performance liquid chromatography (HPLC)) and dissolved in nuclease-free water (Ambion) to 10 mg mL⁻¹. For SAXS experiments, monodispersed Lambda dsDNA (New England BioLabs) or Escherichia coli genomic dsDNA (Affymetrix) was precipitated and resuspended in 100 mM NaCl + 10 mM HEPES (pH 7.4) to 5 mg mL⁻¹. For cell experiments, endotoxin-free human (hdsDNA) and mouse (mdsDNA) genomic self-dsDNA were purchased (Biochain) and used without further purification. Peptide-dsDNA complexes were formed by incubating the peptide with dsDNA (1–5 mg mL⁻¹ for SAXS experiments and 10–20 μg mL⁻¹ for cell experiments) at specific peptide-to-dsDNA charge ratios (P/dsDNA = 1:4, 1:2, 1:1, 2:1, 4:1). For a particular peptide, the same charge ratios were used in both SAXS and cell experiments. For some experiments, complexes were mixed in the presence of human serum (Sigma-Aldrich, H4522).

Levofloxacin (LVFX), N-methylmorpholine (NMM) and resazurin were purchased from Sigma-Aldrich, Co. LLC (St. Louis, MO, USA). Bicinchoninic acid (BCA) and SYTOX Green were purchased from ThermoFisher Scientific, Inc. (Waltham, MA, USA). Fluoro-N,N,N′,N′-tetramethylformamidinium hexafluorophosphate (TFFH) was purchased from EMD Millipore (a division of Merck KGaA; Darmstadt, Germany), Fmoc-Gly-Wang resin from Peptides International, Inc. (Louisville, KY, USA), and DiSC(3)5 from Anaspec, Inc. (Fremont, CA, USA). Deionized water (dH₂O) was prepared using a Milli-Q Synthesis A10 system (Millipore). The bacterial strains E. coli (#25922) and B. cereus (#11778) were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA). Pep-4 (≥95% purity) was custom synthesized by Genscript USA, Inc. (Piscataway, NJ, USA). Melittin (≥95%) was purchased from Anaspec, Inc., and indolicidin was synthesized (≥95% purity) by AAPPTec, LLC (Louisville, KY, USA). Peptide identities were verified via matrix assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry using a Shimadzu AXIMA Performance. Additionally, Pep-4 and the conjugate were submitted for amino acid analysis (UC Davis Genome Center Proteomics Core Facility) in order to establish molar concentrations.