Aurein-3.1

Aurein 2.1, aurein 2.6 and aurein 3.1 are amphibian host defence peptides that kill bacteria via the use of lytic amphiphilic α-helical structures. The C-terminal PEGylation of these peptides led to decreased antibacterial activity (Minimum Lethal Concentration (MLCs) ↓ circa one and a half to threefold), reduced levels of amphiphilic α-helical structure in solvents (α-helicity ↓ circa 15.0%) and lower surface activity (Δπ ↓ > 1.5 mN m-1). This PEGylation of aureins also led to decreased levels of amphiphilic α-helical structure in the presence of anionic membranes and zwitterionic membranes (α-helicity↓ > 10.0%) as well as reduced levels of penetration (Δπ ↓ > 3.0 mN m-1) and lysis (lysis ↓ > 10.0%) of these membranes. Based on these data, it was proposed that the antibacterial action of PEGylated aureins involved the adoption of α-helical structures that promote the lysis of bacterial membranes, but with lower efficacy than their native counterparts. However, PEGylation also reduced the haemolytic activity of native aureins to negligible levels (haemolysis ↓ from circa 10% to 3% or less) and improved their relative therapeutic indices (RTIs ↑ circa three to sixfold). Based on these data, it is proposed that PEGylated aureins possess the potential for therapeutic development; for example, to combat infections due to multi-drug resistant strains of S. aureus, designated as high priority by the World Health Organization.

Aurein 2.6-COOH and aurein 3.1-COOH were studied along with their naturally occurring C-terminally amidated analogues. Circular dichroism (CD) and molecular dynamic (MD) simulations were used to study the effects of amidation on the interaction of antimicrobial peptides (AMPs) with lipid bilayers. CD measurements and MD analysis suggested that both peptide analogues were predominantly random coil and adopted low levels of α-helical structure in solution (<30%) and in the presence of a lipid bilayer the peptides formed a stable α-helical structure. In general, amidated analogues have a greater propensity than the non-amidated peptides to form a α-helical structure. MD simulations predicted that aurein 2.6-COOH and aurein 3.1-CHOOH destabilised lipid bilayers from 1,2-dimyristoyl-sn-glycero-3-phosphocholine and 1,2-dimyristoyl-sn-glycero-3-phosphoserine via angled bilayer penetration. They also showed that aurein 2.6-CONH₂ and aurein 3.1-CONH₂ formed a helix horizontal to the plane of an asymmetric interface.

Seventeen aurein peptides are present in the secretion from the granular dorsal glands of the Green and Golden Bell Frog Litoria aurea, and 16 from the corresponding secretion of the related Southern Bell Frog L. raniformis. Ten of these peptides are common to both species. Thirteen of the aurein peptides show wide-spectrum antibiotic and anticancer activity. These peptides are named in three groups (aureins 1-3) according to their sequences. Amongst the more active peptides are aurein 1.2 (GLFDIIKKIAESF-NH2), aurein 2.2 (GLFDIVKKVVGALGSL-NH2) and aurein 3.1 (GLFDIVKKIAGHIAGSI-NH2). Both L. aurea and L. raniformis have endoproteases that deactivate the major membrane-active aurein peptides by removing residues from both the N- and C-termini of the peptides. The most abundant degradation products have two residues missing from the N-terminal end of the peptide. The solution structure of the basic peptide, aurein 1.2, has been determined by NMR spectroscopy to be an amphipathic alpha-helix with well-defined hydrophilic and hydrophobic regions. Certain of the aurein peptides (e.g. aureins 1.2 and 3.1) show anticancer activity in the NCI test regime, with LC50 values in the 10-5-10-4 M range. The aurein 1 peptides have only 13 amino-acid residues: these are the smallest antibiotic and anticancer active peptides yet reported from an anuran. The longer aurein 4 and 5 peptides, e.g. aurein 4.1 (GLIQTIKEKLKELAGGLVTGIQS-OH) and aurein 5. 1 (GLLDIVTGLLGNLIVDVLKPKTPAS-OH) show neither antibacterial nor anticancer activity.