Generating Biologically Stable TNA Aptamers that Function with High Affinity and Thermal Stability

Abstract

Aptamers are often prone to nuclease digestion, which limits their utility in many biomedical applications. Here we describe a xeno-nucleic acid system based on α-L-threofuranosyl nucleic acid (TNA) that is completely refractory to nuclease digestion. The use of an engineered TNA polymerase permitted the isolation of functional TNA aptamers that bind to HIV reverse transcriptase (HIV RT) with K D 's of ∼0.4−4.0 nM. The aptamers were identified using a display strategy that provides a powerful genotype−phenotype linkage. The TNA aptamers remain active in the presence of nuclease and exhibit markedly higher thermal stability than monoclonal antibodies. The combined properties of biological stability, high binding affinity, and thermal stability make TNA aptamers a powerful system for the development of diagnostic and therapeutic agents.

Metal Nanoparticles for Electrochemical Sensing: Progress and Challenges in the Clinical Transition of Point-of-Care Testing

“…Aptamer spatial configurations are changed to enable the best interaction with the target biomolecules [84]. Additionally, aptamers are more stable than antigens, and can easily recover their active spatial configuration after usage thereby allowing for the reuse of the same electrode multiple times [85]. Based on how the immobilized aptamers interact with the target analyte, Han and coworkers proposed the following categories: (a) spatial configuration rearrangement of aptamers based on target interaction [86][87][88]; (b) sandwich type interactions [89][90][91]; (c) dissociation or displacement of aptamers through target interaction [92,93]; and (d) competitive replacement of aptamers [14,[94][95][96].…”

“…It can be seen from the table that MNPs with sizes between 5 and 20 nm are the most-used individual component in the ECB fabrication process [4,23,26,27,29,41,44]. Although size variation is well studied, the shape of the MNPs used is almost always spherical [4,26,41,44,48,85]. The area of MNP shape control deserves significant attention, because NPs with unique shapes (hollow spheres, cubic, porous, pyramidal, etc.)…”

“…In short, bioaffinity ECB functions are based on such binding capacities of the BRCs via interactions with the nanostructured electrode materials. The BRCs are mainly antibodies (mono/polyclonal), aptamers (single stranded ssDNA sequence/RNA), peptides, etc., which can effectively capture target antigens or biomarkers while constructing bioaffinity ECBs [44,85,98,155]. Table 3 shows the mostly studied bioaffinity ECBs for different biomarkers based on MNPs and nanocomposites assays.…”

In Vitro Selection of an ATP-Binding TNA Aptamer

“…Despite having a backbone repeat unit that is one atom shorter than the repeat unit found in natural DNA and RNA, TNA is capable of forming antiparallel Watson–Crick duplex structures with DNA, RNA, and itself [ 17 , 18 ]. Although TNA aptamers have been generated against several protein targets [ 19 , 20 , 21 ], very little is known about the ability for TNA to recognize small molecule targets with high affinity and specificity [ 22 ]. Here, we report the in vitro selection of an ATP-binding TNA aptamer that shows high specificity against other nucleotide triphosphates and strong resistance to nuclease degradation.…”

Recent advances in aptamer applications for analytical biochemistry

“…This chemical diversity-driver for achieving better performing aptamers has been echoed by Lokesh et al [ 133 ] and Zon [ 28 ]. Recently, such diversity has been increased by Dunn et al [ 134 ] using a novel xeno-nucleic acid (XNA) system in the form α- l -threofuranosyl nucleic acid (TNA) that is completely refractory to nuclease digestion, yet binds to HIV reverse transcriptase with Kd values of ~0.4–4.0 nM. Also, Tolle et al [ 135 ] have extended well-known azide-alkyne cycloaddition-click chemistry [ 136 ] to the solid-phase for high fidelity synthesis of nucleobase-modified DNA aptamers dubbed “clickmers”, which has the potential to greatly expand the chemical diversity of aptamers and, hence, targets for new or improved applications.…”