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Updated: Jan 8, 2026
Identifying the Binding Proteins of Small Ligands with the Differential Radial Capillary Action of Ligand Assay DRaCALA
Published on: March 19, 2021
The solute-binding proteins DppA1-5 of Pseudomonas aeruginosa have distinct substrate profiles
Konstantin Plöchl1,2, Thomas Böttcher3
1Faculty of Chemistry, Institute of Biological Chemistry & Centre for Microbiology and Environmental Systems Science, University of Vienna, Vienna, Austria.
Abstract:
Pseudomonas aeruginosa is an opportunistic pathogen with high antibiotic resistance that infects immunocompromised patients. Its survival in diverse environments relies on nutrient uptake systems such as the dipeptide permease (Dpp), an ATP-binding cassette transporter that imports di- and tripeptides. Unlike other bacteria, whose Dpp includes a single solute-binding protein that binds substrates in the periplasm, P. aeruginosa encodes five paralogs (DppA1-5), the functional significance of which remains unclear. Here, we systematically profile the ligand specificities of all five DppA proteins using Differential Scanning Fluorimetry with a library of 281 di- and tripeptides. We find that DppA1 and DppA3 preferentially bind dipeptides, whereas DppA2 and DppA4 favor tripeptides. DppA5 shows no detectable binding, suggesting a divergent function. Ligand binding is highly structure-sensitive and distinct across DppA paralogs. A comparative reanalysis of published nutrient utilization data for dppA1-5 suggests that previous studies underestimated this functional specialization. In contrast, our findings indicate that P. aeruginosa encodes multiple DppA paralogs to expand Dpp substrate scope and enhance nutrient acquisition. This work provides a foundation for further exploration of the Dpp in bacterial signaling and its exploitation for drug delivery via Trojan Horse antimicrobials.
Insights
Pseudomonas aeruginosa utilizes five dipeptide permease A (DppA) proteins to import various peptides, expanding nutrient acquisition. This functional specialization enhances bacterial survival and offers potential antimicrobial drug delivery strategies.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Pseudomonas aeruginosa is an opportunistic pathogen known for high antibiotic resistance.
- Nutrient uptake systems, like the dipeptide permease (Dpp), are crucial for bacterial survival.
- P. aeruginosa uniquely encodes five paralogs of the Dpp solute-binding protein (DppA1-5), with unclear functional roles.
Purpose of the Study:
- To systematically characterize the ligand specificities of all five P. aeruginosa DppA paralogs.
- To elucidate the functional significance of multiple DppA paralogs in P. aeruginosa.
- To explore the potential of the Dpp system for antimicrobial drug delivery.
Main Methods:
- Differential Scanning Fluorimetry (DSF) was employed to profile ligand specificities.
- A comprehensive library of 281 di- and tripeptides was used for binding assays.
- Comparative reanalysis of existing nutrient utilization data was performed.
Main Results:
- DppA1 and DppA3 preferentially bind dipeptides.
- DppA2 and DppA4 show a preference for tripeptides.
- DppA5 exhibited no detectable binding, indicating a divergent function, and ligand binding was structure-sensitive and paralog-specific.
Conclusions:
- P. aeruginosa possesses functionally specialized DppA paralogs, expanding its peptide substrate uptake range.
- This specialization enhances nutrient acquisition, contributing to bacterial survival in diverse environments.
- Understanding DppA specificity provides a basis for developing Trojan Horse antimicrobials.
