Development of Processes for Recombinant L-Asparaginase II Production by Escherichia coli Bl21 (De3): From Shaker to Bioreactors

Abstract

Since 1961, L-asparaginase has been used to treat patients with acute lymphocytic leukemia. It rapidly depletes the plasma asparagine and deprives the blood cells of this circulating amino acid, essential for the metabolic cycles of cells. In the search for viable alternatives to produce L-asparaginase, this work aimed to produce this enzyme from Escherichia coli in a shaker and in a 3 L bioreactor. Three culture media were tested: defined, semi-defined and complex medium. L-asparaginase activity was quantified using the β-hydroxamate aspartic acid method. The defined medium provided the highest L-asparaginase activity. In induction studies, two inducers, lactose and its analog IPTG, were compared. Lactose was chosen as an inducer for the experiments conducted in the bioreactor due to its natural source, lower cost and lower toxicity. Batch and fed-batch cultures were carried out to reach high cell density and then start the induction. Batch cultivation provided a final cell concentration of 11 g L−1 and fed-batch cultivation produced 69.90 g L−1 of cells, which produced a volumetric activity of 43,954.79 U L−1 after lactose induction. L-asparaginase was produced in a shaker and scaled up to a bioreactor, increasing 23-fold the cell concentration and thus, the enzyme productivity.

ClearColi as a platform for untagged pneumococcal surface protein A production: cultivation strategy, bioreactor culture, and purification

“…There are few studies addressing the influence of inducer (lactose or IPTG) concentration on recombinant protein production by conventional E. coli BL21(DE3) cells under lac operon control (Einsfeldt et al 2011 ; Lecina et al 2013 ; Ferreira et al 2018 ; Barros et al 2021 ; Von den Eichen et al 2021 ), but none for ClearColi® BL21(DE3) so far. Segovia-Trinidad et al ( 2021 ) reported that other induction methods and inducer concentrations did not increase soluble production of constitutive-active human-Smad8, but they did not detail their investigation nor present the results.…”

Produção de L-asparaginase por Pichia pastoris em cultivos descontínuos-alimentados em biorreatores.

“…Em busca de maior produtividade, Barros et al (2021) Sajitha, Vidya, Varsha, Binod, & Pandey (2015), originando outra variante de EcA com glicosilações com padrão mais similar ao de mamíferos.…”