Coupling p-Hydroxybenzoate Hydroaxylase with the Photoresponsive Nanozyme for Universal Dehydrogenase-Based Bioassays

Herein, an innovative dehydrogenase invoked tri-enzyme cascade has been successfully constructed and utilized as a powerful and universal colorimetric strategy for dehydrogenase-based bioassays. Specifically, the glucose-6-phosphatedehydrogenase (G6PD) catalyzes the specific dehydrogenation of glucose-6-phosphate (G-6-P) by consuming the oxidized form of β-nicotinamide adenine dinucleotide phosphate (NADP + ) to generate NADPH. And the formed NADPH initiates the irreversible hydroxylation of p-hydroxybenzoate by p-hydroxybenzoate hydroxylase (PHBH) to generate 3,4-dihydroxybenzoate (DHB), which results in the subsequent in situ generation of a photoresponsive nanozyme of DHB coordinated SrTiO 3 (SrTiO 3 /DHB) nanosheets. The photoresponsive nanozyme has intriguing interfacial charge transfer (ICT) characteristics and thus realizes the efficiently catalytic oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) under light irradiation for signal readout. Exemplified by G6PD and G-6-P as analytes, the dehydrogenase invoked tri-enzyme cascade achieved efficient signal amplification towards the ultrasensitive colorimetric bioassays. This study opens up the ICT transitions of SrTiO 3 for innovative dehydrogenases-involved enzyme cascades. In addition, it is an entirely new method for dehydrogenases-based bioassays, with the distinctive merits of facile operation, rapid response, high sensitivity, and naked-eye readout. Importantly, it is extensible as a general basis for other numerous targets of interest because of the large numbers of dehydrogenases available for catalysing different reactions.