OpenLB
Open Library of Bioscience

The increase in antibiotic-resistant bacterial infections has prompted significant academic research into new therapeutic agents targeted against these pathogens. Antimicrobial peptides (AMPs) appear as promising candidates, due their potent antimicrobial activity and their ubiquitous presence in almost all organisms. tritrpticin is a member of this family of peptides and has been shown to exert a strong antimicrobial activity against several bacterial strains. tritrpticin's main structural characteristic is the presence of three consecutive Trp residues at the center of the peptide. These residues play an important role in the activity of tritrpticin against Escherichia coli. In this work, a recombinant version of tritrpticin was produced in E. coli using calmodulin as a fusion protein expression tag to overcome the toxicity of the peptide. When used in combination with glyphosate, an inhibitor of the endogenous synthesis of aromatic amino acids, this expression system allowed for the incorporation of fluorinated Trp analogs at very high levels (>90%). The antimicrobial activity of the 4-, 5- and 6-fluoro-Trp-containing tritrpticins against E. coli was as strong as the activity of the native peptide. Similarly, the tritrpticin analogs exhibited comparable abilities to perturb and permeabilize synthetic lipid bilayers as well as the outer and inner membrane of E. coli. Furthermore, the use of 19F NMR spectroscopy established that each individual fluoro-Trp residue interacts differently with SDS micelles, supporting the idea that each Trp in the original tritrpticin plays a different role in the perturbing/permeabilizing activity of the peptide. Moreover, our work demonstrates that the use of fluoro-Trp in solvent perturbation 19F NMR experiments provides detailed site-specific information on the insertion of the Trp residues in biological membrane mimetics. This article is part of a Special Issue entitled: Antimicrobial peptides edited by Karl Lohner and Kai Hilpert.