Open AccessExpression of Bacillus amyloliquefaciens transglutaminase in recombinant E. coli under the control of a bicistronic plasmid system in DO-stat fed-batch bioreactor cultivations
Author(s) -
Lovaine Silva Duarte,
Carla Roberta Matte,
Paulo Roberto Dall Cortivo,
J. Nunes,
Laísa Quadros Barsé,
Cristiano Valim Bizarro,
Marco Antônio Záchia Ayub
Publication year - 2021
Publication title -
brazilian journal of microbiology
Language(s) - English
Resource type - Journals
eISSN - 1678-4405
pISSN - 1517-8382
DOI - 10.1007/s42770-021-00521-3
Subject(s) - bacillus amyloliquefaciens , bioreactor , recombinant dna , plasmid , tissue transglutaminase , escherichia coli , enzyme , biology , bacillus (shape) , expression vector , biochemistry , chemistry , microbiology and biotechnology , gene , fermentation , botany
We studied the expression of Bacillus amyloliquefaciens transglutaminase cloned in Escherichia coli BL21(DE3)pLysS harboring the plasmid pBAD/3C/bTGase, a bicistronic expression system, in bioreactor cultivation. Batch and fed-batch controlled as DO-stat strategies were employed for the production of the recombinant enzyme. In 30 h-batch cultivations using Terrific broth (TB), 6 g/L of biomass and 3.12 U/mg protein of transglutaminase activity were obtained. DO-stat fed-batch cultivations under the control of oxygen concentration (DO-stat) using TB as medium but fed with glucose allowed the increment in biomass formation (17.5 g/L) and enzyme activity (6.43 U/mg protein ). DO-stat fed-batch using mineral medium (M9) and fed with glucose under the same conditions produced even higher enzymatic activity (9.14 U/mg protein ). The pH effect was investigated, and the best enzymatic activity could be observed at pH 8. In all cultivations, the bicistronic system remained stable, with 100% of plasmid-bearing cells. These results show that E. coli bearing bicistronic plasmid constructs to express recombinant TGase could be cultivated in bioreactors under DO-stat fed-batch using mineral medium and it is a promising strategy in future optimizations to produce this important enzyme.