IN VITRO SELECTION OF APTAMERS WITH ANTIBACTERIAL POTENTIAL

In vitro selection procedures offer strategies to find novel binders composed of nucleic acids or proteins for a variety of target molecules. DNA and RNA molecules, so called aptamers, have been selected to bind to proteins and peptides, small organic molecules like ATP, antibiotics and even amino acids or inorganic ions with high affinity. In addition, successful selection was carried out to generate aptamers which recognize whole bacterial cells with high specificity.

Here, we applied standard in vitro selection (SELEX) to enrich RNA molecules with binding affinity for the dipeptide L-Ala-L-Ala. The enantiomer D-Ala-D-Ala is part of peptide chains found in the bacterial peptidoglycan layer, where it has the function to crosslink the glycan strands and thereby plays an essential role for the stability of the cell. Hence, enantiomeric aptamers (spiegelmers) with high binding affinity for D-Ala-D-Ala may interfere with the crosslinking process during bacterial growth resulting in an antibacterial effect. Furthermore, these aptamers may be of use for the detection of bacteria.

The initial pool for RNA aptamer selection was generated by partially randomisation of an aptamer selected to bind L-isoleucine with the aim to retrain the aptamer on L-Ala-L-Ala. 15 rounds of selection and amplification resulted in an enrichment of RNA sequences which can be assigned to different sequence families sharing one common motif. Binding tests were carried out to determine binding affinity and specificity. Finally, the antibacterial potential of the spiegelmer form of these aptamers will be analysed.