Design, structural and functional characterization of a Temporin-1b analog active against Gram-negative bacteria

• Published in: Biochimica et biophysica acta Vol. 1830; no. 6; pp. 3767 - 3775
• Main Authors: Avitabile, Concetta, Netti, Fortuna, Orefice, Giuseppina, Palmieri, Maddalena, Nocerino, Nunzia, Malgieri, Gaetano, D'Andrea, Luca D., Capparelli, Rosanna, Fattorusso, Roberto, Romanelli, Alessandra
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.06.2013
• Subjects: Amphibian Proteins - chemical synthesis; Amphibian Proteins - chemistry; Amphibian Proteins - pharmacology; Animals; Anti-Bacterial Agents - chemical synthesis; Anti-Bacterial Agents - chemistry; Anti-Bacterial Agents - pharmacology; anti-infective properties; Antimicrobial activity; antimicrobial peptides; Binding; Drug Design; Escherichia coli; Escherichia coli - growth & development; fluorescence; Gram-negative bacteria; Gram-positive bacteria; hemolysis; LPS; nuclear magnetic resonance spectroscopy; Proteins - chemical synthesis; Proteins - chemistry; Proteins - pharmacology; Rana temporaria; Structure; Temporin; Binding; Antimicrobial activity; Temporin; Structure; LPS
• ISSN: 0304-4165; 0006-3002; 1872-8006
• DOI: 10.1016/j.bbagen.2013.01.026

Temporins are small antimicrobial peptides secreted by the Rana temporaria showing mainly activity against Gram-positive bacteria. However, different members of the temporin family, such as Temporin B, act in synergy also against Gram-negative bacteria. With the aim to develop a peptide with a wide spectrum of antimicrobial activity we designed and analyzed a series of Temporin B analogs. Peptides were initially obtained by Ala scanning on Temporin B sequence; antimicrobial activity tests allowed to identify the TB_G6A sequence, which was further optimized by increasing the peptide positive charge (TB_KKG6A). Interactions of this active peptide with the LPS of E. coli were investigated by CD, fluorescence and NMR. TB_KKG6A is active against Gram-positive and Gram-negative bacteria at low concentrations. The peptide strongly interacts with the LPS of Gram-negative bacteria and folds upon interaction into a kinked helix. Our results show that it is possible to widen the activity spectrum of an antimicrobial peptide by subtle changes of the primary structure. TB_KKG6A, having a simple composition, a broad spectrum of antimicrobial activity and a very low hemolytic activity, is a promising candidate for the design of novel antimicrobial peptides. The activity of antimicrobial peptides is strongly related to the ability of the peptide to interact and break the bacterial membrane. Our studies on TB_KKG6A indicate that efficient interactions with LPS can be achieved when the peptide is not perfectly amphipathic, since this feature seems to help the toroidal pore formation process. [Display omitted] ► A new Temporin B analog, TB_KKG6A, was designed. ► TB_KKG6A is active against Gram-positive and Gram-negative bacteria. ► The peptide strongly interacts with the LPS of E. coli. ► The peptide folds into a kinked helix upon interaction with LPS. ► The peptide shows very low hemolytic activity.