Linearized esculentin-2EM shows pH dependent antibacterial activity with an alkaline optimum

• Published in: Molecular and cellular biochemistry 2021-10, Vol.476 (10), p.3729-3744
• Main Authors: Malik, Erum, Phoenix, David A., Snape, Timothy J., Harris, Frederick, Singh, Jaipaul, Morton, Leslie H. G., Dennison, Sarah R.
• Format: Article
• Language: English
• Subjects: Amphibian Proteins - chemistry; Amphibian Proteins - pharmacology; Analysis; Anti-Bacterial Agents - chemistry; Anti-Bacterial Agents - pharmacology; Antibacterial activity; Antibacterial agents; Antimicrobial Cationic Peptides - chemistry; Antimicrobial Cationic Peptides - pharmacology; Bacteria; Biochemistry; Biomedical and Life Sciences; Cardiology; Disinfection; Ethylenediaminetetraacetic acid; Food; Food preservation; Food spoilage; Gram-negative bacteria; Gram-positive bacteria; Gram-Positive Bacteria - growth & development; Hydrogen-Ion Concentration; Life Sciences; Linearization; Medical Biochemistry; Membranes; Oncology; Peptides; pH effects; Phosphatidylethanolamine; Phosphatidylglycerol; Pore formation; Protein Conformation, alpha-Helical; Spoilage; Theoretical analysis
• ISSN: 0300-8177; 1573-4919
• DOI: 10.1007/s11010-021-04181-7

Here the hypothesis that linearized esculentin 2EM (E2EM-lin) from Glandirana emeljanovi possesses pH dependent activity is investigated. The peptide showed weak activity against Gram-negative bacteria (MLCs ≥ 75.0 μM) but potent efficacy towards Gram-positive bacteria (MLCs ≤ 6.25 μM). E2EM-lin adopted an α-helical structure in the presence of bacterial membranes that increased as pH was increased from 6 to 8 (↑ 15.5–26.9%), whilst similar increases in pH enhanced the ability of the peptide to penetrate (↑ 2.3–5.1 mN m − 1 ) and lyse (↑ 15.1–32.5%) these membranes. Theoretical analysis predicted that this membranolytic mechanism involved a tilted segment, that increased along the α-helical long axis of E2EM-lin (1–23) in the N → C direction, with −   increasing overall from circa  − 0.8 to − 0.3. In combination, these data showed that E2EM-lin killed bacteria via novel mechanisms that were enhanced by alkaline conditions and involved the formation of tilted and membranolytic, α-helical structure. The preference of E2EM-lin for Gram-positive bacteria over Gram-negative organisms was primarily driven by the superior ability of phosphatidylglycerol to induce α-helical structure in the peptide as compared to phosphatidylethanolamine. These data were used to generate a novel pore-forming model for the membranolytic activity of E2EM-lin, which would appear to be the first, major reported instance of pH dependent AMPs with alkaline optima using tilted structure to drive a pore-forming process. It is proposed that E2EM-lin has the potential for development to serve purposes ranging from therapeutic usage, such as chronic wound disinfection, to food preservation by killing food spoilage organisms.