Roles of the Conserved Amino Acid Residues in Reduced Human Defensin 5: Cysteine and Arginine Are Indispensable for Its Antibacterial Action and LPS Neutralization.

Antimicrobial peptides (AMPs) that are able to neutralize toxins are promising antibiotics. In this study we investigated the role of structurally conserved amino acids in reduced human defensin 5 (HD5 ), which is an endogenous peptide with antibacterial action and the ability to neutralize lipopolysaccharide (LPS). Cys residues and high Arg content, rather than Gly and Arg -Glu , were found to be indispensable for HD5 binding to lipid A, for penetrating the bacterial outer and inner membranes, and for eliminating bacteria. Otherwise, all the conserved sites were requisite for HD5 to block the interaction between LPS and LPS-binding protein and to suppress the TLR4-NF-κB signaling pathway initiated by LPS. Accordingly, we designed the acetamidomethylated Cys-E21R-HD5 , which was much more potent than HD5 at eliminating bacteria and which can neutralize LPS. Cys-E21R-HD5 was also found to exhibit a synergistic effect with ciprofloxacin in killing multidrug-resistant Acinetobacter baumannii. The results of this study, in which multifunctional AMPs were designed based on structure-activity research, may help in the development of more peptide antibiotics.