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
