Immobilization of penicillin G acylase using permeabilized Escherichia coli whole cells within chitosan beads

Entrapment of permeabilized whole cells within a matrix is a common method for immobilization. Chitosan possesses distinct chemical and biological properties, which make it a suitable matrix for entrapment and immobilization of penicillin G acylase (PGA). In the first step, Escherichia coli (ATCC 11105) cells were permeabilized using N-cetyl-N,N,N-trimethyl ammonium bromide (CTAB) (0.1% w/v, 45 min, 45 rpm) which then immobilized using glutaraldehyde (5% w/v) as cross-linker and chitosan (3% w/v) as the matrix. These conditions were established after preliminary trials with CTAB and glutaraldehyde concentrations in the range of 0.05-0.25% w/v and 1-9% v/v, respectively. The hydrolytic activity was assayed using Ehrlich reagent. Permeabilization of cells caused 9% increase in Penicillin G Acylase (PGA) conversion after 15 min compared to the intact cells. Although, immobilization on chitosan decreased the conversion compared to un-immobilized treated cells (13%), the new biocatalyst showed acceptable operational stability, maintaining more than 90% of the initial activity after 20 cycles. Optimum conditions for immobilization of E. coli cells were: CTAB 0.1% w/v and glutaraldehyde 5% v/v. A new combination method was successfully developed and optimized for immobilization of treated whole cells on chitosan matrix.