To Fold or Not to Fold: Diastereomeric Optimization of an α‑Helical Antimicrobial Peptide

Membrane disruptive α-helical antimicrobial peptides
(AMPs)
offer an opportunity to address multidrug resistance; however, most
AMPs are toxic and unstable in serum. These limitations can be partly
overcome by introducing D-residues, which often confers protease resistance
and reduces toxicity without affecting antibacterial activity, presumably
due to lowered α-helicity. Here, we investigated 31 diastereomers
of the α-helical AMP KKLLKLLKLLL. Three diastereomers containing
two, three, and four D-residues showed increased antibacterial effects,
comparable hemolysis, reduced toxicity against HEK293 cells, and excellent
serum stability, while another diastereomer with four D-residues additionally
displayed lower hemolysis. X-ray crystallography confirmed that high
or low α-helicity as measured by circular dichroism indicated
α-helical or disordered structures independently of the number
of chirality switched residues. In contrast to previous reports, α-helicity
across diastereomers correlated with both antibacterial activity and
hemolysis and revealed a complex relationship between stereochemistry,
activity, and toxicity, highlighting the potential of diastereomers
for property optimization.