Regenerated coenzyme-based preparation of bienzyme-polymer nanoconjugates and their applications for the synthesis of ethyl (R)-2-hydroxy-4-phenylbutyrate

A modular approach was applied for the synthesis of bienzyme-polymer nanoconjugates (nano-BECs) (50–70 nm) consisting of two enzymes (carbonyl reductase and glucose dehydrogenase) conjugated within a single universal polymer scaffold. The amount of the product ethyl (R)-2-hydroxy-4-phenylbutyrate (R-HPBE) with nano-BECs as the catalyst was 533 mM in a dibutyl phthalate-phosphate buffer (dibutyl phthalate-PB) biphasic system, while the amount of R-HPBE was 349 mM using carbonyl reductase-poly(acrylic acid) as the catalyst, indicating that the nano-BECs have an advantage for coenzyme regeneration. Compared with a single aqueous phase, the substrate treatment capacity was improved at the interface of the dibutyl phthalate-PB biphasic system. Under the optimal reaction conditions (35 °C, 40 h, dibutyl phthalate-PB 1: 1), nano-BECs can completely convert substrate into optically pure R-HPBE (enantiomeric excess (e.e.) >99.9%) in the organic-aqueous system.