Efficient biosynthesis of (R)-1-[3,5-bis(trifluoromethyl)phenyl] ethanamine with self-assembled dual-enzyme clusters

Purified SpyCatcher-ATA117 (5 mu M) and SpyTag-ADH (5 mu M) were covalently combined to form self-assembled dual-enzyme clusters (DENCs) at 4 degrees C for 3 h. The successful assembly of DENCs was verified using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), transmission electron microscopy (TEM), scanning electron microscopy (SEM), and dynamic light scattering (DLS). DENCs improved the storage stability by 36.2% and pH stability by 11.2% compared with those of free double enzymes. 56.89 g/L (R)- 1-[3,5-bis (trifluoromethyl)phenyl]ethanamine (R-BPA) was synthesized using DENCs with high yield (99.9%) and high enantiomeric excess (99.9%) at 45 degrees C and pH 9 for 24 h. Moreover, DENCs retained more than 78% of the initial activity after being reused in the preparation of R-BPA for five cycles. Thus, DENCs were successfully formed, and efficient amination was achieved, providing a new biocatalyst for efficient biosynthesis of chiral amines via asymmetric transamination cascade reaction.

Automated summary

In this study, self-assembled dual-enzyme clusters (DENCs) were successfully prepared and their assembly was verified. DENCs showed improved storage stability and pH stability compared to free enzymes. High-yield (99.9%) and high-enantiomeric excess (99.9%) synthesis of R-BPA was achieved using DENCs, and the DENCs retained high activity after five cycles.