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Updated: Oct 10, 2025
Antimicrobial Peptides Produced by Selective Pressure Incorporation of Non-canonical Amino Acids
Published on: May 4, 2018
Effect of Amino Acid Substitutions on 70S Ribosomal Binding, Cellular Uptake, and Antimicrobial Activity of Oncocin
Lisa Kolano1,2, Daniel Knappe1,3, Angela Berg4
1Institute of Bioanalytical Chemistry, Faculty of Chemistry and Mineralogy, 04103, Leipzig, Germany.
Abstract:
Proline-rich antimicrobial peptides (PrAMPs) are promising candidates for the treatment of infections caused by high-priority human pathogens. Their mode of action consists of (I) passive diffusion across the outer membrane, (II) active transport through the inner membrane, and (III) inhibition of protein biosynthesis by blocking the exit tunnel of the 70S ribosome. We tested whether in vitro data on ribosomal binding and bacterial uptake could predict the antibacterial activity of PrAMPs against Gram-negative and Gram-positive bacteria. Ribosomal binding and bacterial uptake rates were measured for 47 derivatives of PrAMP Onc112 and compared to the minimal inhibitory concentrations (MIC) of each peptide. Ribosomal binding was evaluated for ribosome extracts from four Gram-negative bacteria. Bacterial uptake was assessed by quantifying each peptide in the supernatants of bacterial cultures. Oncocin analogues with a higher net positive charge appeared to be more active, although their ribosome binding and uptake rates were not necessarily better than for Onc112. The data suggest a complex mode of action influenced by further factors improving or reducing the antibacterial activity, including diffusion through membranes, transport mechanism, secondary targets, off-target binding, intracellular distribution, and membrane effects. Relying only on in vitro binding and uptake data may not be sufficient for the rational development of more active analogues.
Insights
Proline-rich antimicrobial peptides (PrAMPs) show potential against pathogens. However, in vitro binding and uptake data alone do not fully predict their antibacterial activity, suggesting a complex mechanism of action.
Area of Science:
- Antimicrobial Peptides
- Molecular Biology
- Drug Development
Background:
- Proline-rich antimicrobial peptides (PrAMPs) are investigated for treating infections from high-priority pathogens.
- PrAMPs act via membrane diffusion, transport, and inhibition of protein biosynthesis by targeting the 70S ribosome exit tunnel.
Purpose of the Study:
- To determine if in vitro ribosomal binding and bacterial uptake data can predict the antibacterial activity of PrAMPs.
- To evaluate 47 derivatives of the PrAMP Onc112 against Gram-negative and Gram-positive bacteria.
Main Methods:
- Measured ribosomal binding of PrAMPs to ribosome extracts from four Gram-negative bacteria.
- Assessed bacterial uptake by quantifying peptide concentrations in bacterial culture supernatants.
- Compared these in vitro measures to the minimal inhibitory concentrations (MIC) of each peptide.
Main Results:
- Oncocin analogues with higher net positive charge showed increased activity.
- Higher net positive charge did not consistently correlate with improved ribosome binding or uptake rates compared to Onc112.
- Antibacterial activity appears influenced by multiple factors beyond simple binding and uptake.
Conclusions:
- In vitro data on ribosomal binding and bacterial uptake are insufficient for predicting the full antibacterial spectrum of PrAMPs.
- The mode of action is complex, involving factors like membrane diffusion, transport, secondary targets, and intracellular distribution.
- Rational development of novel PrAMP analogues requires consideration of these additional complex factors.
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