Encapsulation of recombinant Escherichia coli cells harboring nitrilase in surface-crosslinked hydrogel particles: A robust and durable biocatalyst for the hydrolysis of nitrile

Abstract

Surface-crosslinked hydrogel particles encapsulating E. coli cells harboring nitrilase were constructed and used as a durable biocatalyst in the synthesis of gabapentin intermediate. E. coli cells (50 g/L) expressing nitrilase were encapsulated in 2% (w/v) alginate hydrogel, 0.5%(w/v) glutaraldehyde and 1%(w/v) polyethyleneimine were sequentially used as crosslinker. The constructed alginate particles with GA&PEI composite crosslinking had a similar catalytic capacity compared to equivalent free cells, which could completely hydrolyze 1.0 M substrate 1-cyanocyclohexaneacetonitrile within 4 h at pH 7.5 and 35 °C, and have a specific activity of 134.6 U/g, an optimum temperature at 55 °C and a corresponding half-life up to 19.1 ± 1.9 h. The particles showed excellent storage stability, reusability and mechanical strength, which retained 90.3% activity after 2-month storage at 4 °C, and 83.2% activity after 14-day storage at 35 °C. In batch reactions, 50 g/L particles could be reused for maximum 32 cycles, maintained 63.5% residual activity and 50.7% particle integrity, providing an average product yield of 94.6% in the first 30 batches and a productivity of 1467.03 g/L/day. Besides, the stability and catalytic activity of the particles were almost unaffected in several organic co-solvents and in purified water, showing their good industrial potential.