Amplified Biosensing Using the Horseradish Peroxidase-Mimicking DNAzyme as an Electrocatalyst

Abstract

The hemin/G-quadruplex horseradish peroxidase-mimicking DNAzyme is assembled on Au electrodes. It reveals bioelectrocatalytic properties and electrocatalyzes the reduction of H(2)O(2). The bioelectrocatalytic functions of the hemin/G-quadruplex DNAzyme are used to develop electrochemical sensors that follow the activity of glucose oxidase and biosensors for the detection of DNA or low-molecular-weight substrates (adenosine monophosphate, AMP). Hairpin nucleic structures that include the G-quadruplex sequence in a caged configuration and the nucleic acid sequence complementary to the analyte DNA, or the aptamer sequence for AMP, are immobilized on Au-electrode surfaces. In the presence of the DNA analyte, or AMP, the hairpin structures are opened, and the hemin/G-quadruplex horseradish peroxidase-mimicking DNAzyme structures are generated on the electrode surfaces. The bioelectrocatalytic cathodic currents generated by the functionalized electrodes, upon the electrochemical reduction of H(2)O(2), provide a quantitative measure for the detection of the target analytes. The DNA target was analyzed with a detection limit of 1 x 10(-12) M, while the detection limit for analyzing AMP was 1 x 10(-6) M. Methods to regenerate the sensing surfaces are presented.

Guanine Quadruplex Electrochemical Aptasensors

“…Since the first report on using hemin/G4 DNAzyme as the electrocatalytic label for amplifying sensing events [102], this approach attracted increasing interest in biosensor [103,104] and biofuel cell technologies [105]. In comparison with protein peroxidases, the hemin/G4 peroxidase-mimicking DNAzymes have several advantages, such as high chemical stability, low cost and simple synthesis.…”

“…In comparison with protein peroxidases, the hemin/G4 peroxidase-mimicking DNAzymes have several advantages, such as high chemical stability, low cost and simple synthesis. Hemin/G4 DNAzyme electrochemical biosensors were successfully used for the detection of cells [106,107], proteins [108][109][110] or low molecular weight molecules, such as adenosine monophosphate (AMP) [102,111,112], anticancer drugs [113], gaseous ligands [114], toxins [115,116], pollutant agents [117,118] or metal ions [119,120].…”

“…The glucose oxidase activity was followed by a hemin/G4 DNAzyme electrochemical biosensor, by attaching the enzyme to the electrode surface through the nucleic acid sequence able to form G4s in the presence of hemin [102,124]. Then, the glucose oxidase mediated the glucose oxidation to gluconic acid and H 2 O 2 , and the resulting H 2 O 2 was analyzed through its electrocatalyzed reduction by the DNAzyme.…”

Optical and electrochemical detection of a verotoxigenic E. coli gene using DNAzyme-labeled stem-loops

“…On the other hand, electrochemical transduction has been chosen when DNA strands were immobilized onto electroactive surfaces [10][11][12]. From the biosensors construction viewpoint, electrochemical detection shows a higher degree of electronic integration and a simplified design resulting from the fact that an electric signal is the natural result of electrochemical transduction [13].…”

DNA sensing by electrocatalysis with hemoglobin

“…Enzymes such as horseradish peroxide and glucose oxidase have been shown to readily interact electronically with phenothiazine dyes, such as MB, and have had wide application as recognition elements for molecular sensing (33)(34)(35)(36)(37)(38)(39)(40)(41)(42). Detection platforms based on these enzymes are desirable due to their inherent catalytic activity.…”