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J App Pharm Sci. 2017; 7(8): 149-155


Bench-scale Overproduction and Purification of recombinant GCSF in Escherichia coli fed-batch process

Valiollah Babaeipour; Mohammad Reza Mofid; Sirwan Khanchezar; Faezeh Faraji; Somayeh Abolghasemi.

Abstract
Fermentation process scale up can change the hydrodynamic conditions of recombinant E. coli cultivation bioreactor and reduce productivity in the same operating conditions that have been developed at lab scale. In this study, an efficient strategy was used to scale up recombinant human GCSF production in the fed-batch culture of E. coli. Heterologous production of human GCSF was done in the fed-batch culture of Escherichia coli BL21 (DE3) with feeding strategy of maximum attainable specific growth rate. Then, fed-batch process was scaled up based on geometrical analogy and constant DO. Rh-GCSF was overexpressed as inclusion body (IB). Then IBs were released by cell disruption afterward solubilized and refolded properly. Finally, the rh-GCSF was purified by cation exchanger chromatography. By induction at cell density of 75 g dry cell weight per l (g dcw/l), final cell density and rh-GCSF concentration in both scales were 124 and 114 g dcw/l, and 22 and 19 g protein/l obtained, respectively. Recovery yield of purification process was 400 mg purified gcsf per 1 g IB and purity of recombinant protein was over than 99%. Insignificant decrease of the biomass and rh-GCSF production (less than 10 percent) during scale up indicates efficiency of scale up method.

Key words: : rh-GCSF;Fed-Batch;Geometrical Analogy;Overproduction;Purification;Escherichia coli


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