Antimicrobial and Antibiofilm Activities of Helical Antimicrobial Peptide Sequences Incorporating Metal-Binding Motifs

Abstract

Antimicrobial peptides (AMPs) represent alternative strategies to combat the global health problem of antibiotic resistance. However, naturally occurring AMPs are generally not sufficiently active for use as antibiotics. Optimized synthetic versions incorporating additional design principles are needed. Here, we engineered amino-terminal Cu(II) and Ni(II) (ATCUN) binding motifs, which can enhance biological function, into the native sequence of two AMPs, CM15 and citropin1.1. The incorporation of metalbinding motifs modulated the antimicrobial activity of synthetic peptides against a panel of carbapenem-resistant enterococci (CRE) bacteria, including carbapenem-resistant Klebsiella pneumoniae (KpC+) and Escherichia coli (KpC+). Activity modulation depended on the type of ATCUN variant utilized. Membrane permeability assays revealed that the in silico selected lead template, CM15, and its ATCUN analogs increased bacterial cell death. Mass spectrometry, circular dichroism, and molecular dynamics simulations indicated that coordinating ATCUN derivatives with Cu(II) ions did not increase the helical tendencies of the AMPs. CM15 ATCUN variants, when combined with Meropenem, streptomycin, or chloramphenicol, showed synergistic effects against E. coli (KpC+ 1812446) biofilms. Motif addition also reduced the hemolytic activity of the wild-type AMP and improved the survival rate of mice in a systemic infection model. The dependence of these bioactivities on the particular amino acids of the ATCUN motif highlights the possible use of size, charge, and hydrophobicity to fine-tune AMP biological function. Our data indicate that incorporating metal-binding motifs into peptide sequences leads to synthetic variants with modified biological properties. These principles may be applied to augment the activities of other peptide sequences.

Antimicrobial Peptides: Classification, Design, Application and Research Progress in Multiple Fields

“…Besides, the Cu-ATCUN derivative of OV-3 containing a C-terminal GGC sequence showed high levels of membrane permeation and lipid peroxidation. The concept of catalytic metal drugs has attracted widespread attention although the concept is still in its infancy because of the role of metal ions (Alexander et al, 2017;Agbale et al, 2019).…”

Antimicrobial Susceptibility Testing of Antimicrobial Peptides to Better Predict Efficacy

“…Transition metal ions influence the activity of AMP in a variety of ways. In some cases, the activity of AMP are dependent on the presence of metal ions (Paulmann et al, 2012 ; Melino et al, 2014 ; Alexander et al, 2018 ; Jezowska-Bojczuk and Stokowa-Soltys, 2018 ; Agbale et al, 2019 ), whereas in others the presence of metal ions can cause reduction, or even complete abrogation, of activity (Friedrich et al, 1999 ; Deslouches et al, 2005 ). In most circumstances, the effect of metal ions must be considered on a case-by-case basis as different ions will produce a distinct effect.…”

Copper-binding motifs Xxx-His or Xxx-Zzz-His (ATCUN) linked to an antimicrobial peptide: Cu-binding, antimicrobial activity and ROS production

“…However, generally modest effects were observed, limited to the same order of magnitude as the AMP without Cu. Another approach was to add a Cu(II)-binding motif to AMPs by elongating the AMP with a XZH motif [8,[20][21][22][23][24]. In these cases an increased activity by a Cu-ATCUN moiety in AMPs was observed for AMPs proceeding through different mechanisms, suggesting a role for Cu-ATCUN motif not restricted to a single antimicrobial mechanism.…”