Colorimetric Logic Gate for Pyrophosphate and Pyrophosphatase via Regulating the Catalytic Capability of Horseradish Peroxidase

By regulating the catalytic capability of horseradish peroxidase (HRP), an artful colorimetric assay platform for pyrophosphate (PPi) and pyrophosphatase (PPase) was unprecedentedly designed. In this work, Cu(I), generated by reducing Cu(II) in the presence of ascorbate, could inhibit HRP's catalytic capability of transforming colorless 3,3',5,5'-tetramethylbenzidine (TMB) into blue oxidized TMB (oxTMB). The robust coordination between PPi and Cu(II) is able to discourage the reduction of Cu(II) to Cu(I) effectively, thus restoring the original catalytic capability of HRP and regenerating blue-colored oxTMB. Upon PPase introduction, PPi would be hydrolyzed into orthophosphate, which could release Cu(II) free from the Cu(II)-PPi complex, and thus in turn allows the catalytic capability of HRP to be inhibited by Cu(I). HRP was activated or deactivated to different degrees depending on PPi or PPase levels, which could be indicated by using HRP-triggered catalytic system as a signal amplifier, thus paving a way for PPi and PPase sensing. Based on the colorimetric sensor for PPi and PPase, an "INH" logic gate was rationally constructed. With the merits of high sensitivity and selectivity, cost-effectiveness, and simplification, our proposed analytical system has also been verified to have potential to be utilized for enzyme inhibitor screening and diagnosis of PPase-related diseases.