Composite of Si-tag-fused heterologous enzymes and mesoporous silica for the efficient synthesis of an optically active alcohol

Abstract

Efficient synthesis of high-value-added optically active alcohol ((R)-1-phenyl-1,2-ethanediol [(R)-PED]), used as pharmaceutical intermediate, was achieved by coupled enzyme reactions using mesoporous silica as the immobilization scaffold for heterologous enzymes. When heterologous enzymes, (R)-carbonyl reductase (RCR), and sorbitol dehydrogenase (SDH), were immobilized in ordered pores of mesoporous silica, SDH activity in coenzyme regeneration was significantly reduced. To solve this, we fused Si-tag, a silica-binding protein, to both heterologous enzymes (RCR/SDH) and attempted to orientationally immobilize them on the silica surface, leading to significant increase in (R)-PED yield. The yield increased approximately 5 times, from 8–10 % in heterologous enzymes without Si-tag to 40 % in Si-tag-fused heterologous enzymes. While enzyme activity remained low when Si-tag-fused heterologous enzymes were immobilized on non-porous silica support, repeated durability of Si-tag-fused heterologous enzymes immobilized on mesoporous silica improved significantly by expanding pore diameter to 10.6 nm. The (R)-PED yield after repeated use for 10 times was 50 % compared to the first reaction. To improve manageability of the enzyme support, a pelletized mesoporous silica powder–organic polymer composite was prepared. When Si-tag-fused heterologous enzymes immobilized on this pelletized composite were applied to (R)-PED synthesis, high enzyme activity and repeated durability, equivalent to mesoporous silica powder, were exhibited.