Efficient production of (R)-(−)-mandelic acid by isopropanol-permeabilized recombinant E. coli cells expressing Alcaligenes sp. nitrilase

• The catalytic activity of recombinant E. coli cells was increased by 3.6-fold by isopropanol permeabilization.
• The process efficiency of the permeabilized cells-catalyzed reaction was 2-fold that of the non-permeabilized cells.
• The immobilized permeabilized cells could be reused for 15 rounds without significant activity loss.

Resting cells or immobilized cells are preferentially used as biocatalyst for biocatalytic processes due to the easy preparation and operation, however, the catalytic efficiency of biocatalyst is usually limited by the transport barrier of cell membrane. In this work, the transport barrier of recombinant Escherichia coli cells expressing the nitrilase from Alcaligenes sp. ECU0401 was relieved by isopropanol permeabilization. Under the optimal permeabilization conditions, the nitrilase activity of the permeabilized cells was 4.6-fold that of the cells without permeabilization. The permeabilized cells were evaluated for the enantioselective hydrolysis of mandelonitrile, which showed an initial reaction rate of about 2-fold that of the cells without permeabilization. The permeabilization process did not impair the stability of the cells and after simple immobilization by glutaraldehyde cross-linking, the immobilized permeabilized cells were successfully recycled for 15 runs without significant activity loss, suggesting the potential application of the immobilized permeabilized cells for the production of (R)-(−)-mandelic acid. The technology reported in this work might also be extended to other bioprocesses facing cell membrane transport barriers.