Co‐encapsulating Cofactor and Enzymes in Hydrogen‐Bonded Organic Frameworks for Multienzyme Cascade Reactions with Cofactor Recycling

Use of hydrogen‐bonded organic frameworks (HOFs) for enzyme immobilization faces challenges in improving enzyme activity recovery and assembling cofactor‐dependent multienzyme systems. Herein, we report a polyelectrolyte‐assisted encapsulation approach (PAEA) that enables two cascades with four oxidoreductases and two nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) cofactors co‐encapsulated in BioHOF‐1 with excellent cargo loading and over 100% cascade activities. The key role of the polyelectrolyte is to coat enzymes and tether NAD(P)H, thus interacting with HOF monomers in place of enzymes, avoiding enzymes from destruction by HOF monomers. The versatility and efficiency of PAEA are further illustrated by an HOF‐101‐based bio‐nanoreactor. Moreover, the immobilization by PAEA makes enzymes and NAD(P)H display excellent stability and recyclability. This work has demonstrated a facile and versatile PAEA for fabricating cofactor‐dependent multienzyme cascade nanoreactors with HOFs.