Effect of Environmental Conditions on the Expression Levels of a Recombinant 11S Amaranth Globulin in Escherichia coli
Hypatia Arano-Varela, Jorge Dominguez-Dominguez and Octavio Paredes-Lopez
Affiliation: Cinvestav-IPN, Irapuato Unit, Km 9.6 Libramiento Norte Carr. Irapuato-Leon, 36821, Irapuato, Gto., Mexico
Keywords: E. coli-expressed protein, protein ACM3, batch fermentation, environmental conditions, response surface methodology.
The expression in Escherichia coli strain Rosetta of the recombinant acidic subunit from the 11S amaranth seed
storage protein (protein ACM3) was studied at flask and at bioreactor levels. This subunit was modified by inserting four
Val-Tyr antihypertensive peptides in tandem into its third variable region and also with the tripeptide Ile-Pro-Pro in the Cterminal
region. Flasks experiments allowed us to define the best conditions for the preparation and expression and accumulation
of the protein ACM3, including the certainty of its presence within the cells especially as an insoluble fraction.
The effects of cultivation temperature, aeration rate and agitation speed on the production of the protein ACM3 was tested
in a 5-L batch bioreactor. Applying response surface methodology (RSM) we found that the aeration rate was the most
significant factor affecting in a positively way the production yields and productivity of the recombinant protein. Temperature
had effect only in conjunction to aeration. The highest recombinant acidic subunit concentration (747 mg L-1) and
the highest productivity (186 mg L-1 h-1) were attained in 4 h of cultivation when the factors evaluated were controlled at
its central values: 0.1 vvm, 300 rpm, and 30.5°C. Results from this study indicate that RSM is an effective technique to
maximize the production of this recombinant protein.
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