Operation of packed-bed immobilized cell reactor featuring active β-galactosidase inclusion body-containing recombinant Escherichia coli cells

Abstract

In this study, we developed a packed-bed immobilized cell reactor containing active β-gal (β-galactosidase) inclusion body (IB)-containing Escherichia coli (E. coli) cells in alginate beads. This packed-bed reactor was operated using a substrate feed solution 0.72 ∼ 38.4 mM ONPG (o-nitrophenyl-β-D-galactoside) prepared in Z buffer supplemented with chloroform and 0.1% SDS (sodium dodecyl sulfate). The production rate of ONP (o-nitrophenol) in the reactor containing cells that were incubated with α-MG (α-methyl D-glucospyranoside) or D-fucose after induction was superior to those prepared with cells that were not incubated with α-MG or D-fucose. The ONP production rate was increased proportionally with ONPG concentration in the substrate feed up to a concentration of 38.4 mM. However, as the ONPG concentration was increased in the substrate feed solution, galactose inhibition inside the alginate beads was increased. This most likely occurred due to problems with diffusion. In addition, partial breakage of alginate beads was observed during the later periods of operation. In this study, we demonstrated that active β-gal IB-containing E. coli cells were sustained in the immobilized cell reactor during operation. Particularly, these findings demonstrate the feasibility of using active IBs in an enzymatic reaction without the need for any purification step. In addition, we showed that these IB-containing cells could be directly used in an immobilized reactor.