Optimized upstream and downstream process conditions for the improved production of recombinant human asparaginase (rhASP) from Escherichia coli and its characterization

Publication date: Available online 13 October 2018Source: BiologicalsAuthor(s): Rajesh Kumar Kante, Sandeep Vemula, Silpa Somavarapu, Maheswara Reddy Mallu, B.H. Boje Gowd, Srinivasa Reddy RondaAbstractThe present work elucidates the production of recombinant human asparaginase (rhASP) under optimized fermentation and downstream processes in Escherichia coli. The maximum biomass yield of 6.7 g/L was achieved with fed-batch fermentation. The highest rhASP inclusion bodies recovery yield (91%) was achieved with the optimized lysis conditions. The 8.0 M urea at pH 8.5 has shown efficient solubilization (94%) of rhASP inclusion bodies. The refolding efficiency of rhASP increased at pH 8.5 (84%) and temperature 25°C (86%). The diluted rhASP solution was concentrated and partially purified (92%) using cross flow filtration. A single step ion exchange chromatography is successfully achieved the maximum purity of ≥ 97%. The molecular mass of purified rhASP is confirmed as 34.1 kDa by mass spectrometry. The secondary structure of rhASP is characterized by FT-IR spectroscopy based on the structural elements. Finally, cell proliferative assay of purified rhASP is signifies the similar biological activity over the standard.

Source: Biologicals - October 13, 2018 Category: Biology Source Type: research