A new method for the in situ assay of α-glucosidase activity and inhibitor screening through an enzyme substrate-mediated DNA hybridization chain reaction

Abstract

In this work, a new strategy has been proposed for the in situ analysis of α-glucosidase (α-Glu) based on the small molecule-mediated DNA hybridization chain reaction (HCR). As an important digestion enzyme, α-Glu could catalyze the hydrolysis of the substrate para-aminophenyl-α-D-glucopyranoside (pAPG) into para-aminophenyl (pAP) and α-D-glucopyranoside. Among them, pAPG with cis-diols groups could coordinate with the boronic acid-tagged initiation chain to drive HCR around the assembled magnetic nanospheres (AMNSs). Nevertheless, the product of the catalytic reaction pAP did not have the capability to capture the boronic acid-tagged initiation chain onto the surface of AMNSs, due to which HCR did not occur. Thus, the α-Glu activity was correlated with the extent of HCR around AMNSs and it could be tested in situ in a cell medium with the aid of a magnet. The enzyme activity was analyzed over a wide linear range from 0.4 U mL⁻¹ to 1.7 U mL⁻¹ with the lowest detection limit of 0.023 U mL⁻¹. Furthermore, the established method could also be successfully utilized for the in situ evaluation of enzyme inhibitors including gallic acid and quercetin. This study may provide a new insight for the development of a new method for the in situ analysis of α-Glu in a cell medium.