Highly Efficient Biosynthesis of Nicotinic Acid by Immobilized Whole Cells of E. coli Expressing Nitrilase in Semi-Continuous Packed-Bed Bioreactor

A recombinant E. coli, expressing nitrilase from Acidovorax facilis 72W with dual-site expression plasmid pRSFduet (E. coli pRSF-AfNit2), was constructed. It showed higher soluble of than that in the pET21a plasmid. The can efficiently catalyze hydrolysis 3-cyanopyridine to nicotinic acid. whole cells pRSF-AfNit2 were immobilized by using sodium alginate/glutaraldehyde/polyethylene imine as best reagents. 95% activity recovery and excellent mechanical strength, improved thermal stability pH stability. They also retained 82% initial after nearly two months storage at 4 °C. semi-continuous packed-bed bioreactor (sPBR) filled studied for efficient production After optimization, highest space–time yield 1576 g/(L·d) obtained on 0.8 M substrate concentration 2 mL/min flow rate. sPBR repeatedly operated 41 batches, keeping 100% conversion presence 30 mM CaCl2. Finally, 95 g acid 90% separation purification. developed technology has potential application value.