madman
Super Moderator
pg.13 (pdf)
Aptamers are more present now than ever in scientific research.
Since their discovery in 1990 [1,2], aptamers have piqued the interest of scientists looking for bioreceptors for small molecules. Numerous aptamers, capable of binding their small molecule target with high affinity and selectivity are described in literature. They can be implemented into biosensors by using a fairly simple and logical design that links a target induced conformational change of the aptamer to a signal generation that can be ‘read’ electrochemically or by fluorescence. Although a number of these biosensors appear successful, many challenges remain to be addressed for these to break-through in real world applications [3]. One challenge concerns addressing the lack of insight at the molecular level of the factors determining affinity and selectivity, as well as going beyond the simplified, cartoon-like approach in describing the underlying sensing mechanism. We make an effort to contribute towards this goal by developing NMR based strategies, using the structure switching testosterone binding TESS.1 DNA aptamer [4] as model system.
To verify interaction between the TESS.1 aptamer and its said target testosterone, we analysed and identified its corresponding complexation patterns in different regions of the proton NMR spectrum. To allow improved analysis and interpretation of the aptamer target interaction, a two-step optimisation was performed, generating a more ‘NMR optimal’ construct, which we labelled TESS.1_s_mod. It interacts with the target in a similar fashion as the originally sized TESS.1 aptamer. A more complete assignment of the system was secured by introducing single nucleotide 13C and 15N uniform labelling of G and T nucleotides in TESS.1_s_mod. Besides a more in depth analysis of binding events, these assignments will also allow 3D structure determination.
We will present and discuss these developed NMR based strategies and recent results that provide the first detailed molecular view on the TESS.1 aptamer and its interaction with the target testosterone.
Figure 1: The proposed 2D representation, based on [4], of the TESS.1 aptamer and its target testosterone.
www.ncbi.nlm.nih.gov
Aptamers are more present now than ever in scientific research.
Since their discovery in 1990 [1,2], aptamers have piqued the interest of scientists looking for bioreceptors for small molecules. Numerous aptamers, capable of binding their small molecule target with high affinity and selectivity are described in literature. They can be implemented into biosensors by using a fairly simple and logical design that links a target induced conformational change of the aptamer to a signal generation that can be ‘read’ electrochemically or by fluorescence. Although a number of these biosensors appear successful, many challenges remain to be addressed for these to break-through in real world applications [3]. One challenge concerns addressing the lack of insight at the molecular level of the factors determining affinity and selectivity, as well as going beyond the simplified, cartoon-like approach in describing the underlying sensing mechanism. We make an effort to contribute towards this goal by developing NMR based strategies, using the structure switching testosterone binding TESS.1 DNA aptamer [4] as model system.
To verify interaction between the TESS.1 aptamer and its said target testosterone, we analysed and identified its corresponding complexation patterns in different regions of the proton NMR spectrum. To allow improved analysis and interpretation of the aptamer target interaction, a two-step optimisation was performed, generating a more ‘NMR optimal’ construct, which we labelled TESS.1_s_mod. It interacts with the target in a similar fashion as the originally sized TESS.1 aptamer. A more complete assignment of the system was secured by introducing single nucleotide 13C and 15N uniform labelling of G and T nucleotides in TESS.1_s_mod. Besides a more in depth analysis of binding events, these assignments will also allow 3D structure determination.
We will present and discuss these developed NMR based strategies and recent results that provide the first detailed molecular view on the TESS.1 aptamer and its interaction with the target testosterone.
Figure 1: The proposed 2D representation, based on [4], of the TESS.1 aptamer and its target testosterone.
High-Affinity Nucleic-Acid-Based Receptors for Steroids - PMC
Artificial receptors for hydrophobic molecules usually have moderate affinities and limited selectivities. We describe three new classes of high affinity hydrophobic receptors for non-aromatic steroids based on deoxyribonucleotides, obtained through ...
www.ncbi.nlm.nih.gov
