Aptamer‐Assisted Bioconjugation of Tyrosine Derivatives with hemin/G‐quadruplex (hGQ) DNAzyme Nucleoapzyme Nanostructures

Abstract

Hemin/G-quadruplex (hGQ) DNAzymes are horseradish peroxidase-mimicking catalysts capable of the oxidation of a variety of substrates. We now implement aptamer-functionalized hGQ DNAzymes, also known as nucleoapzymes, to achieve increased bioconjugation of N-methyl luminol to tyrosinecontaining residues and peptides. We found that the presence of a tyrosinamide-binding aptamer leads to a 12-fold increase in the catalytic rate constant (k cat ), and the saturation kinetics curves that were obtained provide evidence for the involve-ment of the substrate binding site in the reaction. The application of the best performing nucleoapzymes for the modification of Tyr-containing peptides reveals that (i) the aptamer also recognizes the ligand structure when this is embedded in a larger peptide structure, and (ii) distant residues in the peptide substrate can influence the conversion. As such, we show that nucleoapzymes display enzyme-like features and provide an additional tool in the toolbox of bioconjugation chemistry.

Functionalized oligonucleotides, synthetic catalysts as enzyme mimics

“…An application of this concept was described for the bioconjugation of tyrosine derivatives using N-methyl luminol. The impact of this aptamer-assisted catalysis could be demonstrated by the 12-fold enhancement observed when the aptamer sequence binds to the tyrosine substrate [47].…”

DNA nanocrane-based catalysts for region-specific protein modification

“…[1][2][3][4][5] Although finely-tuned chemical agents and precisely tailored catalytic constructs have yielded approaches that enable the modification of a selective subset of identical functional groups, 2,3,[6][7][8][9] precise control over modification of native proteins remains challenging, especially within a complex biological setting such as a cell. Our past studies were directed at the development of DNAzymes as enzyme mimics for protein modification, [10][11][12] the enrichment of thrombin-binding aptamer with catalytic moieties that lead to trigger-responsive site-selective and protein-selective modification, 12 and calibration of the modification efficiency and range with respect to the distance between catalyst and protein. 13 So far, however, it remained to be seen whether complex DNA-based catalytic nanostructures would also be able to selectively modify a protein of interest (POI) in the presence of many other proteins.…”

Catalytic DNA Nanocranes for Controlled Protein Modification in Cell Lysate

“…[1][2][3][4][5] Although finely-tuned chemical agents and precisely tailored catalytic constructs have afforded approaches that enable the modification of a selective subset of identical functional groups, 2,3,[6][7][8][9] precise control over modification of native proteins remains challenging. Our past studies were directed at the development of DNAzymes as enzyme mimics for protein modification, [10][11][12] the enrichment of thrombin-binding aptamer with catalytic moieties that lead to trigger-responsive site-selective and proteinselective modification, 12 and calibration of the modification efficiency and range with respect to the distance between catalyst and protein. 13 In this study, we seek to integrate various elements into novel catalytic structures that (i) reversibly interact with the target protein, (ii) enable control over the protein region that is subjected to modification, and (iii) allow this modification to be performed in the complex mixture of a cell lysate.…”