Cascade-Amplified Time-Resolved Fluorescent Assay Driven by an Enzyme-Integrated Catalytic Compartment as an Artificial Multi-Enzyme Complex.

We here report a simple and efficient strategy of fabricating artificial multi-enzyme complex (MEC) based on the integration of natural enzyme with catalytic compartment. As a proof of concept, this strategy was demonstrated by selecting cholesterol oxidase (ChOx) and CeIII -based nanoscale coordination polymer (Ce-NCP) with peroxidase-like activity as the models, which forms ChOx@Ce-NCP. Benefitting from the confinement and sheltering effects of Ce-NCP, superior cascade activity and stability in harsh environments were achieved in ChOx@Ce-NCP. Meanwhile, the distinct advantage of ChOx@Ce-NCP has also been highlighted by its negligible substrate inhibition effect and adjustable mass ratio of building blocks. Upon the doping of TbIII in ChOx@Ce-NCP, a luminescent artificial MEC (ChOx@Ce-NCP:Tb) was further fabricated to drive a cascade amplified time-resolved fluorescent assay within a confined space, showing high sensitivity and specificity toward cholesterol.