Multifunctional G‐Quadruplex Aptamers and Their Application to Protein Detection

Two significant G-quadruplex aptamers named AGRO100 and T30695 are identified as multifunctional aptamers that can bind the protein ligands nucleolin or HIV-1 integrase and hemin. Besides their strong binding to target proteins, both AGRO100 and T30695 exhibit high hemin-binding affinities comparable to that of the known aptamer (termed PS2M) selected by the in vitro evolution process. Most importantly, their corresponding hemin-DNA complexes reveal excellent peroxidase-like activities, higher than that of the reported hemin-PS2M DNAzyme. This enables these multifunctional aptamers to be applied to the sensitive detection of proteins, which is demonstrated by applying AGRO100 to the chemiluminescence detection of nucleolin expressed at the surface of HeLa cells. Based on the specific AGRO100-nucleolin interaction, the surface-expressed nucleolin of HeLa cells is labeled in situ with the hemin-AGRO100 DNAzyme, and then determined in the luminol-H(2)O(2) system. Through this approach, the sensitive detection of total nucleolin expressed at the surface of about 6000 HeLa cells is accomplished. Our results suggest that exploiting new functions of existing aptamers will help to extend their potential applications in the biochemical field.