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Phthalic Acid Ester-Binding DNA Aptamer Selection, Characterization, and Application to an Electrochemical Aptasensor
Published on: March 21, 2018
Pyoverdine binding aptamers and label-free electrochemical detection of pseudomonads
Sharif Anisuzzaman1,2, Nima Alimoradi3, Dilini Singappuli-Arachchige1,4
1Ames Laboratory, U. S. Department of Energy, Ames, IA, United States.
Abstract:
Pyoverdines are iron-chelating siderophores employed by various pseudomonads to promote their growth in iron-limited environments, facilitating both beneficial and detrimental interactions with co-inhabiting microbes or hosts, including plants and animals. The fluorescent pseudomonads produce fluorescent pyoverdines comprised of a conserved central chromophore and a unique strain-specific peptidic side chain produced by non-ribosomal peptide synthetases. Pyoverdine Pf5 (PVD-Pf5) is produced by Pseudomonas protegens Pf-5, a species known for supporting plant growth and its involvement in plant pathogen control. To develop a means of exploring the dynamics of P. protegens activity in soil and in the rhizosphere, we selected DNA aptamers that specifically recognize PVD-Pf5 with high affinities. Two selected aptamers with only 16% identity in sequence were examined for structure and function. We found evidence that both aptamers form structures in their apo-forms and one aptamer has structural features suggesting the presence of a G-quadruplex. Although their tertiary structures are predicted to be different, both aptamers bind the target PVD-Pf5 with similar affinities and do not bind other siderophores, including the related pyoverdine, pseudobactin, produced by Pseudomonas sp. B10. One aptamer binds the pyoverdine peptide component and may also interact with the chromophore. This aptamer was integrated into a nanoporous aluminum oxide biosensor and demonstrated to successfully detect PVD-Pf5 and not to detect other siderophores that do not bind to the aptamer when evaluated in solution. This sensor provides a future opportunity to track the locations of P. protegens around plant roots and to monitor PVD-Pf5 production and movement through the soil.
Insights
Researchers developed DNA aptamers to detect Pyoverdine Pf5 (PVD-Pf5), a siderophore from Pseudomonas protegens. This innovation enables tracking P. protegens in soil and monitoring PVD-Pf5 production near plant roots.
Area of Science:
- Microbiology
- Biochemistry
- Biosensor Technology
Background:
- Pyoverdines are iron-chelating siderophores produced by pseudomonads, crucial for microbial interactions in various environments.
- Pseudomonas protegens Pf-5 produces Pyoverdine Pf5 (PVD-Pf5), influencing plant growth and pathogen control.
- Understanding P. protegens dynamics in soil requires methods to track its activity and PVD-Pf5 production.
Purpose of the Study:
- To develop specific DNA aptamers for recognizing and detecting PVD-Pf5.
- To create a biosensor for monitoring PVD-Pf5 in environmental samples.
- To facilitate the study of P. protegens in soil and rhizosphere.
Main Methods:
- Selection of high-affinity DNA aptamers against PVD-Pf5.
- Structural analysis of selected aptamers, including G-quadruplex investigation.
- Integration of a PVD-Pf5-specific aptamer into a nanoporous aluminum oxide biosensor.
- Testing biosensor specificity against other siderophores.
Main Results:
- Two DNA aptamers with high affinity and specificity for PVD-Pf5 were identified.
- One aptamer showed potential interaction with both the peptide and chromophore components of PVD-Pf5.
- A PVD-Pf5-detecting biosensor was successfully developed using one aptamer.
- The biosensor demonstrated specificity, detecting PVD-Pf5 while ignoring other siderophores.
Conclusions:
- DNA aptamers can be effectively designed to target specific siderophores like PVD-Pf5.
- Aptamer-based biosensors offer a promising tool for environmental monitoring of microbial metabolites.
- This technology can advance research on P. protegens ecological roles and interactions.

