Structure-guided design of a Z_basic2-mediated dual-enzyme nanoreactor for chiral amine synthesis

The synthesis of chiral amines is of critical importance but still challenging. Here, we present a self-sufficient and reusable dual-enzyme nanoreactor for chiral amine synthesis, featuring Z_basic2-mediated site-specific immobilization of amine dehydrogenase (AmDH) and glucose dehydrogenase (GDH) onto mesoporous silica nanoflowers (MSN). Molecular dynamics simulations revealed that the Z_basic2 tag was bound to MSN via electrostatic interactions, thus maintaining the fusion enzyme's active pocket accessibility and improving its catalytic performance. Using the Z_basic2 tags, AmDH and GDH were purified and immobilized on MSN in a one-pot process, thus creating the dual-enzyme nanoreactor. The dual-enzyme system exhibited remarkable activity and reusability, achieving high yields of 76-99 % (ee > 99 %) for various chiral amines at a substrate concentration of 400 mM and retaining a yield of 65 % after 10 cycles. This Z_basic2-based platform offers a robust and scalable strategy for sustainable chiral amine synthesis.