Development of an Industrially Feasible Medium for Enhanced Production of Extremely Thermophilic Recombinant Endo-l,4-β-xylanase by Escherichia coli

摘要

Escherichia coli BL21 (DE3) with a plasmid vector pET-22b (+) carrying xylanase coding gene isolated from an extremely thermophilic bacterium, Thermotoga neapolitana was used in current study for the production of recombinant endo-1,4-β-xylanase (EC 3.2.1.8) which is also called xylanase in short. The study was focused on development of an industrially feasible production media for enhanced production of xylanase. Plackett–Burman based model was initially applied to identify the significant media components effecting the xylanase production, followed by a linear full factorial experimental design (General Linear Model), to estimate the optimum concentration ranges of those significant components and the interactions between them. Finally, Box-Behnken based design was applied to fine tune the concentration rages of significant media components as well as the inducer. Using response maximizing tool, up to 878.72 IU mL~(-1 ) of xylnase activity was predicted in media composition (in g L~(-1)): Maltose, 22.8; (NH_(4))H_(2)PO_(4,)10.5; K_(2)HPO_(4), 16.5; KH_(2)PO_(4), 3.6; MgSO_(4).7H_(2)O, 2; and trace element solution (TES), 3.8 mL.L~(-1) when induced with 8g.L~(-1) of lactose at mid log phase of cell growth. During the predicted model validation, the intracellular xylanase production was found enhanced up to of 800 IU mL~(-1 ) which was around 3 folds increase in productivity when compared to IPTG induced production in unoptimized medium. As a part of scaling up the process, the optimized media composition and cultivation conditions were then tested in pilot scale bioreactor and up to 5600IU mL~(-1 ) of xylnase activity was achieved at the end of batch cultivation.