Poly-Arginine Tails and Helical Segments of Natural Antimicrobial Peptides Display Concerted Action at Membranes for Enhanced Antimicrobial Effects

Abstract

Sequence motifs or patterns found in natural antimicrobial peptides (AMPs) have a great impact on their microbicidal activities. Here, through database inquiries and biological assays, we explore the enhanced antibacterial function associated with poly arginine (poly-R) motifs that typically occur as 3–5 concatenated R residues in many natural AMPs. Using a suite of biophysical techniques, we explore the structural consequences of a C-terminal poly-R motif at membranes and correlate our findings with the functional assays. We use natural peptides, such as Tilapia piscidin 4 (TP4), as an example of how various segments in an AMP play separate and synergistic roles to achieve unmatched bactericidal effects. The function of the poly-R segment is highly consequential since the simple addition of a five-arginine (R5) tail to an otherwise inert and weakly binding helical peptide creates a potent AMP. We investigate interactions of AMPs with lipid bilayers mimicking bacterial membrane compositions, including lipopolysaccharides, to show that the poly-R tail has a key role in initiating membrane destabilization through lipid segregation and water sequestration effects, all of which facilitate insertion and translocation of the amphipathic, α-helical N-terminal segment through the membrane. We compiled a large set of natural AMP sequences and MIC values to show that, statistically, the poly-R sequence motifs have, in average, a greater impact on the overall antimicrobial efficacy than equivalent sequences with poly-K motifs and similar charge densities. We discuss our observations in light of the unique structural and hydration properties of arginine residues.