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

ABSTRACT

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 (HD5RED ), which is an endogenous peptide with antibacterial action and the ability to neutralize lipopolysaccharide (LPS). Cys residues and high Arg content, rather than Gly18 and Arg6 -Glu14 , were found to be indispensable for HD5RED binding to lipid A, for penetrating the bacterial outer and inner membranes, and for eliminating bacteria. Otherwise, all the conserved sites were requisite for HD5RED 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 Acm Cys-E21R-HD5RED , which was much more potent than HD5RED at eliminating bacteria and which can neutralize LPS. Acm Cys-E21R-HD5RED 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.