Improved β-Glucanase Production by a RecombinantEscherichia coli Strain using Zinc-Ion Supplemented Medium

In order to investigate the suitability of different metal chelates for affinity chromatography, an expression vector was constructed. It contained a hybrid beta-glucanase as a model protein fused with a His(6)-tag and a secretion cassette providing the ability to secrete beta-glucanase into the culture medium. Supplementation of zinc to the medium led to a rapidly increased expression and release of the target protein into the cultivation medium. Results in respect to the supplementation of the commonly used Terrific Broth "TB-medium" with different metal ions are reported with special emphasis on the influence of zinc ions. A concentration of zinc ions in the order of about 0.175 mM led to optimal results. Batch cultivation under well-controlled conditions showed that the growth behavior did not change significantly by adding zinc ions. Growth in a stirred tank bioreactor was much faster in unsupplemented TB-medium compared to shake flask experiments leading to a much higher biomass concentration (15 g/L instead of 3 g/L). The secretion of beta-glucanase under theses conditions started at the transition into the stationary phase and increased to yield an extracellular activity of 1350 U/mL at the end of the fermentation process. An even higher yield of extracellular beta-glucanase (2800 U/mL) was reached when the fermentation was carried out with TB-medium supplemented with 0.175 mM ZnSO4.