Structure-Function Analysis of Tritrypticin, an Antibacterial Peptide of Innate Immune Origin

Abstract

The structural requirements for the antibacterial activity of a pseudosymmetric 13-residue peptide, tritrypticin, were analyzed by combining pattern recognition in protein structures, the structure-activity knowledgebase, and circular dichroism. The structure-activity analysis, based on various deletion analogs, led to the identification of two minimal functional peptides, which by themselves exhibit adequate antibacterial activity against Escherichia coli and Salmonella typhimurium. The common features between these two peptides are that they both share an aromatic-proline-aromatic (ArProAr) sequence motif, and their sequences are retro with respect to one another. The pattern searches in protein structure data base using the ArProAr motif led to the identification of two distinct conformational clusters, namely polyproline type II and ␤-turn, which correspond to the observed solution structures of the two minimal functional analogs. The role of different residues in structure and function of tritrypticin was delineated by analyzing antibacterial activity and circular dichroism spectra of various designed analogs. Three main results arise from this study. First, the ArProAr sequence motif in proteins has definitive conformational features associated with it. Second, the two minimal bioactive domains of tritrypticin have entirely different structures while having equivalent activities. Third, tritrypticin has a ␤-turn conformation in solution, but the functionally relevant conformation of this gene-encoded peptide antibiotic may be an extended polyproline type II.

Rationale-Based, De Novo Design of Dehydrophenylalanine-Containing Antibiotic Peptides and Systematic Modification in Sequence for Enhanced Potency

“…Four lysine residues in such an arrangement were expected to form a polar face, while ⌬Phe residues align themselves to form a hydrophobic face, resulting in an amphipathic helical structure. A tryptophan residue was included in the design because of its known membrane-active properties and association with antibacterial activity in peptides like indolicidin and tritrypticin (24,36). Peptides varying from 10 to 12 residues have shown significant antimicrobial activity (15).…”

Ion channel-like activity of the antimicrobial peptide tritrpticin in planar lipid bilayers

“…Several studies aimed at understanding the mechanism of action of Trp3 peptide in lipid membranes have been performed [8,[10][11][12][13]. However, no clear description has been obtained thus far.…”

Cationic Antimicrobial Peptides

“…Examples are: the hexapeptide lactoferricin [58], indolicidin [59], TP [60], Pac-525 [61], the short bovine bactenecins analogs [54], and synthetic linear RW-rich hexapeptides indentified from combinatorial library studies [62]. On the level of the lipid membrane, the W residue has a high propensity to insert into the membrane and partition near the membrane-water interface [63], while positively charged R residues, with hydrogen bonding properties, provide the basis for peptide interaction with the anionic components of bacterial membranes.…”