A DNA functionalized porphyrinic metal-organic framework as a peroxidase mimicking catalyst for amperometric determination of the activity of T4 polynucleotide kinase.

An electrochemical method is described for the sensitive detection of the activity of the enzyme T4 polynucleotide kinase (PNK) by using a DNA functionalized porphyrinic metal-organic framework (L/(Fe-P)n-MOF). In the presence of PNK, the hairpin oligonucleotide (HP1) becomes phosphorylated, and the trigger is released by lambda exonuclease (λ exo). The trigger DNA hybridizes with hairpin probe (immobilized on the gold electrode) to form a nicking endonuclease cleavage site. Thus, a single-strand capture probe is employed to hybridize with L/(Fe-P)n-MOF. The (Fe-P)n-MOF is a peroxidase mimicking material with high catalytic efficiency. By using this amplification strategy, an electrochemical signal is procured that allows for the determination of T4 PNK in the 1.0 mU·mL to 1.0 U·mL with a detection limit of 0.62 mU·mL. The method is selective and can be used to screen for enzyme inhibitors. Conceivably, the (Fe-P)n-MOF can also be used to detect other analytes via its peroxidase-mimicking activity. Graphical abstract Schematic presentation of T4 polynucleotide kinase (PNK) detection. Two hairpin DNAs (HP) and a porphyrinic metal-organic framework with peroxidase-mimicking activity are used. The detection limit is 0.62 mU mL with enzyme assisted signal amplification. This method is selective and can be used to screen for enzyme inhibitors.